US4994443A - Inhibition of angiogenesis - Google Patents

Inhibition of angiogenesis Download PDF

Info

Publication number
US4994443A
US4994443A US07/257,409 US25740988A US4994443A US 4994443 A US4994443 A US 4994443A US 25740988 A US25740988 A US 25740988A US 4994443 A US4994443 A US 4994443A
Authority
US
United States
Prior art keywords
heparin
active agents
cortisone
hexasaccharide
oral
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Fee Related
Application number
US07/257,409
Inventor
Moses J. Folkman
Stephanie Taylor
Robert S. Langer
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Childrens Medical Center Corp
Original Assignee
Childrens Medical Center Corp
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
US case filed in Court of Appeals for the Federal Circuit litigation Critical https://portal.unifiedpatents.com/litigation/Court%20of%20Appeals%20for%20the%20Federal%20Circuit/case/2021-2335 Source: Court of Appeals for the Federal Circuit Jurisdiction: Court of Appeals for the Federal Circuit "Unified Patents Litigation Data" by Unified Patents is licensed under a Creative Commons Attribution 4.0 International License.
US case filed in New Jersey District Court litigation https://portal.unifiedpatents.com/litigation/New%20Jersey%20District%20Court/case/2%3A19-cv-15343 Source: District Court Jurisdiction: New Jersey District Court "Unified Patents Litigation Data" by Unified Patents is licensed under a Creative Commons Attribution 4.0 International License.
Application filed by Childrens Medical Center Corp filed Critical Childrens Medical Center Corp
Priority to US07/257,409 priority Critical patent/US4994443A/en
Application granted granted Critical
Publication of US4994443A publication Critical patent/US4994443A/en
Anticipated expiration legal-status Critical
Expired - Fee Related legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters

Definitions

  • This invention relates to inhibition of angiogenesis and pertains more specifically to treatment of mammals with heparin or heparin fragments or analogues and with steroids to inhibit angiogenesis with subsequent regression of large tumor masses and prevention of tumor metastasis in mammals containing such tumors.
  • Angiogenesis the growth of new capillary blood vessels, is important in normal processes such as development of the embryo, formation of the corpus luteum and wound healing. It is also a component in pathologic processes such as chronic inflammation, certain immune responses and neoplasia. Furthermore, angiogenesis is a property of most solid tumors and is necessary for their continued growth.
  • Heparin an ⁇ , ⁇ glycosidically linked highly sulfated copolymer of uronic acid and glucosamine, has been used clinically as an anticoagulant for half a century. Despite its importance and widespread use, both the exact structure of heparin and the precise nature by which it acts in blood anticoagulation have not been elucidated. Much of the difficulty in determining the structure of heparin is because it is not a homogeneous substance. Heparin is polydisperse with a molecular weight range from 5,000 to 40,000. Within a given chain, there are also structural variations such as varying degrees of sulfation, N-acetylation, and C-5 epimerization in the uronic acid residue.
  • Heparin has been degraded by treatment with heparinase (an enzyme of bacterial origin, Langer et al. U.S. Pat. No.
  • angiogenesis in mammals is inhibited and tumor masses in mammals are caused to regress (and metastasis prevented) by administration of the combination of two essential active agents: (1) heparin or a heparin fragment which is a hexasaccharide or larger, or an analogous compound having one of the structures ##STR1## and (2) a steroid having 17 ⁇ - and 21-hydroxy groups, 3- and 20-one groups, and in the 16- position hydrogen hydroxy or a methyl group, and non-toxic physiologically acceptable carboxylates, acetals, ketals and phosphates thereof, said combination exhibiting a vascular zone when implanted in an immature chick chorioallantoic membrane.
  • steroids which lack glucocorticoid and mineralo-corticoid activity, since such activity is an undesired side effect and limits the dose size or extent of use of the steroid for the purpose of the present invention.
  • steroids are 11 ⁇ , 17,21-trihydroxypregn-4-ene-3,20-dione (or 11 ⁇ -hydrocortisone), 17 ⁇ ,21-dihydroxypregn-4-ene-3,20-dione (or 11-desoxycortisol or Cortexolone), and 17 ⁇ ,21-dihydroxypregna-4,9(11)-diene-3,20-dione.
  • Inhibition of angiogenesis in accordance with the present invention in addition to its effect upon tumor regression and metastasis in tumor-bearing mammals, is effective as a contraceptive in females even if first administered after insemination has occurred, and is effective in treating diseases involving neovascularization such as neovascular diseases of the eye.
  • the active agents may be mixed together prior to administration or may be administered separately at about the same time so that both are present simultaneously in the mammal being treated.
  • the administration may be oral or parenteral including inter alia topical application, intravenous, intra-arterial or subcutaneous injection, and including absorption as well as injection and introduction into bodily apertures or orifices
  • heparin which is commercial available in the form of heparin sodium
  • oral administration leads to degradation in the gastrointestinal tract which results in loss of its anticoagulant activity, but because it has been found that the degradation products include disaccharide and larger fragments, this mode of administration is highly effective for the present invention both for heparin for heparin fragments, and for the specified compounds.
  • heparin and its fragments as well as the specified compounds may be employed in any physiologically acceptable non-toxic form, including their metal salts, preferably as the sodium salts, all of which are embraced in the term "heparin” or “fragment” or “compound” as used in the present specification and claims.
  • metal salts preferably as the sodium salts
  • heparin sold under the trade name "Panheprin” (Abbott Laboratories) is preferred, but heparin from other sources, such as Hepar, Inc. can also be used although less effective
  • Cortisone and its physiologically acceptable non-toxic derivatives such as the acetate, as well as many other steroids useful in the present invention, are only very slightly soluble in water, hence are preferably administered parenterally, e.g. subcutaneously, not orally.
  • steroids such as hydrocortisone or its 11- ⁇ isomer (which are relatively water soluble as compared to cortisone) or one of their water-soluble physiologically acceptable non-toxic derivatives such as a carboxylate, acetal, ketal or the phosphate are preferred
  • Water-insoluble derivatives of water-soluble steroids such as derivatives of hydrocortisone or its 11- ⁇ isomer which are non-toxic and physiologically acceptable, are administered parenterally.
  • the terms "cortisone” and “hydrocortisone” and 11- ⁇ isomer of hydrocortisone as used in the present specification and claims are intended to include both the steroids themselves and their derivatives as defined above.
  • Dosages employed are limited only by the well known limits for the administration of the drugs individually for their usual effects, in the case of cortisone, hydrocortisone, or its 11- ⁇ isomer. However, a number of the useful steroids have no other biological effect apart from the present invention. Simple testing, for example, by the procedure of
  • Heparin may be administered percutaneously in amounts as large as tolerable without objectionable anticoagulant effects. Since heparin administered orally has no anticoagulant effect, and since the hexasaccharide fragment has no anticoagulant effect whether given orally or in any other way, large dosages can thus be administered without risk of bleeding. Oral dosages of heparin of the order of 27,000-45,000 units per kg. body weight per day have been found to be effective, but when administered subcutaneously, doses greater than about 600 units per kg. body weight twice daily led to undesirable anticoagulation effects. In the case of the hexasaccharide fragment, 7 mg per kg. body weight twice daily subcutaneously has been found effective.
  • Cortisone acetate was effective in subcutaneous dosages of 250 mg/kg/day down to 37 mg/kg/day and hydrocortisone was effective orally in amounts of 0.45 mg/ml drinking water (approximately 75 mg/kg/day).
  • the 11- ⁇ isomer of hydrocortisone is approximately equal to hydrocortisone in activity for the purpose of the present invention.
  • the dose size required to bring about regression of tumors or to prevent metastasis varies to some extent depending upon the identity of the tumor, as does the length of time required to bring about complete regression of tumors. Tumor size at the beginning of treatment also affects the length of time required for complete regression. Because of the occurrence of angiogenesis in psoriasis and arthritis, it is expected that the present invention may be useful in treating these diseases. Because administration of cortisone, with or without heparin or heparin fragments or the specified compounds, may result in pulmonary infection after a number of days, it is desirable to administer a suitable antibiotic as prophylaxis during treatment in accordance with the present invention.
  • the active agents are best dissolved or suspended in a suitable carrier which itself must be non-toxic and physiologically acceptable, such as water or normal saline.
  • a suitable carrier which itself must be non-toxic and physiologically acceptable, such as water or normal saline.
  • Compositions containing mixtures of the active agents, either dry or in a suitable carrier, can be employed.
  • Cortisone acetate 0.9 mg in 0.9 ml saline was flooded over the chorioallantoic membrane of 8-day chick embryos through a window in the shell made previously.
  • tumor extract 100 ⁇ g
  • 5 ⁇ l H 2 O was placed on the center of a round 15 mm diameter plastic coverslip and allowed to dry.
  • To the center of each coverslip was then added a 5 ⁇ l aliquot containing either heparin (6 ⁇ g, i.e., 1 unit), or water, at least twenty embryos being subjected to each.
  • the coverslip was placed on the chorioallantoic membrane: Additional control embryos received tumor extract and/or heparin, but were not pre-treated with cortisone acetate. The membranes were viewed on day 11 with a ⁇ 12 stereoscope. Angiogenesis was present if new capillaries were seen to converge on the spot in the center of the coverslip. All of the embryos treated with water or heparin but no cortisone exhibited angiogenesis, as well as 80% of those treated with cortisone acetate alone. Less than 2% of those treated with both heparin and cortisone acetate exhibited angiogenesis.
  • Porcine mucosal heparin was exhaustively degraded using heparinase by the procedure of Langer et al , Science Vol. 217, 261-3 (1982) and the products were fractionated using Sephadex columns equilibrated with 1M NH 4 OAc.
  • the degraded heparin had no anticoagulant activity as determined by activated partial thromboplastic time or whole blood recalcification time.
  • the product or product mixture 250 mg was dissolved in 1 cc of 1M NH 4 OAc, loaded onto a 75 ⁇ 2.5 cm G-15 column, and eluted at 0.5 cc/min.
  • the mixture of tetra-, hexa-, and oligosaccharides was freeze-dried, redissolved in 1 cc of 1M NH OAc, and eluted from a 50 ⁇ 1.25 cm G-50 at 2 cc/min, resulting in an unresolved double peak corresponding to tetra- and hexasaccharide fragments and an additional peak corresponding to oligosaccharides which was freeze-dried.
  • the tetra- and hexasaccharide fragments were combined, freeze-dried, redissolved in 1 cc of 1M NH 4 OAc, and loaded onto a G-15 column.
  • the tetrasaccharide was eluted from a G-15 column in a single peak, the center cut of which was freeze-dried.
  • the hexasaccharide fraction was freeze-dried, redissolved in 0.3 cc 1M NH 4 OAc, eluted from a G-15 column in a single peak, the center cut of which was freeze-dried.
  • heparin fragments were dissolved into methylcellulose discs either alone or with cortisone acetate. The discs were then applied to the 4-day yolk sac membrane of chick embryos cultured in Petri dishes as described by Taylor and Folkman, Nature, Vol. 297, 307-312 (1982). In the presence of cortisone acetate (100 ⁇ g), as shown in the following table, the hexasaccharide fragment demonstrated the highest antiangiogenesis activity.
  • Tetra- and disaccharides were inactive. Oligosaccharides were less active and were toxic at higher concentrations. Therefore, the hexasaccharide fragment was used in subsequent experiments.
  • discs containing hexasaccharide (12 ⁇ g) and cortisone acetate (100 ⁇ g) produced large avascular zones up to 12.6 ⁇ 0.1 mm diameter by 48 hours.
  • cortisone acetate As in the case of heparin (with cortisone acetate), capillaries in the mesodermal layer were absent while the other two tissue layers of the membrane were intact and viable. Hexasaccharide alone did not promote tumor angiogenesis as heparin did.
  • Fertilized chick embryos were removed from their shell on day 3 (or 4) and incubated in a Petri dish in high humidity and 5% CO 2 as previously described by Auerbach et al., J. Devel. Biol., Vol. 41, 391-4 (1974), except that an outer dish and antibiotics were not used.
  • a methylcellulose disc (10 ⁇ l) containing either heparin (6 ⁇ g), or hexasaccharide heparin fragment (12 ⁇ g), or cortisone acetate (Sigma, powder free of preservatives and suspending agents), or a combination of cortisone acetate+heparin or cortisone acetate+hexasaccharide was implanted on the chorioallantoic membrane.
  • the embryos were examined 48 hours later, and if a clear avascular zone appeared around the methylcellulose disc, the diameter of the zone was measured with a Nikon Profile projector at ⁇ 20. Thirty embryos were used in each group. India ink was injected into the heart of some embryos just before formalin fixation so that vessels could be followed to the edge of the avascular zone in histological sections.
  • Hexasaccharide+cortisone acetate produced avascular zones of 12.6 ⁇ 0.1 mm diameter in all embryos. Heparin+cortisone acetate produced avascular zones of 8.9 ⁇ 0.7 mm diameter. There were no avascular zones in the presence of any compounds alone, or with methylcellulose alone.
  • EVA ethylene vinyl acetate copolymer
  • Polymer pellets of ethylene vinyl acetate copolymer (EVA) of approximately 1 mm diameter were impregnated, using the procedure of Langer et al., Nature, Vol. 263, 797-800 (1976), with heparin 180 ⁇ g (Sigma), or hexasaccharide fragment 300 ⁇ g, or cortisone acetate 1.5 mg (Sigma), or a combination of cortisone and heparin.
  • the pellets were implanted in the cornea of a rabbit eye 1 mm from the limbus and a 1 mm 3 piece of V2 carcinoma was implanted distal to the polymer, 2 mm from the limbus. In the opposite eye of each rabbit, control pellets that were empty were similarly implanted in juxtaposition to the tumor.
  • New capillary blood vessels were observed growing towards the tumor and passing over an empty pellet or a pellet containing heparin alone at a mean rate of 0.44 mm/day; and over a pellet containing cortisone alone at 0.22 mm/day.
  • the tumors behind these pellets were vascularized by 6-8 days.
  • the pellets contained both cortisone and heparin there was no capillary growth for 13 days.
  • the heparin-cortisone pellets were removed or when the pellets were depleted of heparin, capillary growth resumed. Histologic sections showed that tumor cells remained viable and capable of replication even when they were adjacent to the heparin-cortisone pellet.
  • Reticulum cell Sarcoma 1 mm 3 pieces of tumor were implanted with a trocar subcutaneously in the backs of 30 mice; 5 per group. Treatment was begun 10 days later, when mean tumor volume was 1.5 ⁇ 10 2 mm 3 .
  • Oral heparin was 200 U/ml in drinking water, average daily consumption was 5-10 ml water per 22 g mouse.
  • Cortisone acetate was administered subcutaneously once daily in a dose of 250 mg/kg for six days, 125 mg/kg for one day, 75 mg/kg for one day, and thereafter a daily maintenance dose of 37 mg/kg (a "tapered" dosage). Control animals received either saline injections, or heparin alone or cortisone acetate alone. All controls were dead by day 34 with large primary tumors and lung metastases. All mice treated with oral heparin+cortisone tapered dosage became tumor-free by day 15 and remained so after treatment was discontinued.
  • mice An additional group of mice was treated similarly except that heparin was administered twice daily subcutaneously in a dose of 627 units, and cortisone acetate was administered subcutaneously once daily in a uniform dose of 75 mg/kg.
  • Response of the mice was the same as in the first set except that tumors recurred after cessation of treatment; these mice became permanently tumor-free when subjected to the regimen of oral heparin and cortisone acetate tapered dosage described above.
  • One of these mice died on day 31 with no gross primary tumor and no metastases.
  • heparin was administered with hydrocortisone, dexamethasone, or medroxyprogesterone. Only hydrocortisone was as effective as cortisone acetate in causing tumor regression when administered with heparin. At the highest tolerable doses neither dexamethasone (3.2 mg/Kg), nor medroxyprogesterone (112 mg/Kg), caused regression of Lewis lung tumors with or without heparin.
  • B-16 Melanoma 7.4 ⁇ 10 6 melanoma cells were injected subcutaneously into 40 mice; 5 per group. Treatment of one group was with oral heparin as described above and oral hydrocortisone, 0.45 mg/ml in drinking water Another group received oral heparin and subcutaneous cortisone acetate tapered dosage, and a third group received subcutaneous heparin and cortisone acetate 75 mg/kg. Controls received either water, or heparin alone, or hydrocortisone or cortisone acetate alone. All control animals died by day 31 with large tumors and lung metastases. In the heparin+cortisone acetate groups, treatment was discontinued after tumor had become invisible for approximately 7 days.
  • Bladder Carcinoma 70 mice: 7 per group received a 1 mm 3 implant of tumor subcutaneously. All control animals died by day 31, with large primary tumors. No mice bearing bladder carcinoma developed lung metastases.
  • the "dormant" tumors were viable, as evidenced by resumption of tumor growth whenever treatment was discontinued for one mouse at a time, beginning at day 70.
  • the hexasaccharide heparin fragment of Example 2 was dissolved in saline, 1.5 mg/ml.
  • Three mice bearing implanted ovarian sarcoma were treated with subcutaneous injection of the hexasaccharide fragment twice daily at a dosage of 7 mg/kg and subcutaneous injection of cortisone acetate- tapered dosage.
  • Control mice received either cortisone acetate alone or saline. While control tumors grew progressively, the hexasaccharide+cortisone acetate treated tumors regressed rapidly and were barely visible 4 days later. Their treatment with hexasaccharide was then discontinued, and the tumors reappeared 3-5 days later.
  • Heparin (oral)-cortisone acetate tapered dosage significantly inhibited capillary growth (0.02 mm/day) compared to cortisone acetate alone (0.24 mm/day), heparin alone (0.32 mm/day) or saline (0.23 mm/day).
  • a thin plate of tumor remained avascular. Three-dimensional tumor-growth did not occur.
  • tumors became vascularized and grew as a three-dimensional mass until they eventually perforated the cornea. These large tumors could be regressed to the flat, thin intracorneal phase by the resumption of the heparin-cortisone acetate combination. However, the intracorneal tumor cells could not be eradicated; discontinuation of the heparin-cortisone acetate led to recurrence of a vascularized tumor.
  • tumor regression might be caused by direct cytotoxicity
  • all 4 types of tumor cells were cultured in the presence of 10% serum obtained from mice receiving either heparin, cortisone acetate, heparin-cortisone acetate, or no drug.
  • Heparin-cortisone acetate did not inhibit cell growth, but in fact stimulated it.
  • histological sections shows no evidence of a cytotoxic effect on bone marrow or intestinal mucosa in animals receiving heparin-cortisone acetate.
  • mice were inoculated with fresh tumor cells at various intervals after they were off treatment. These tumors grew at the same rate as the original implants. Furthermore, if tumor regression was nearly complete and heparin-cortisone acetate was discontinued prematurely, the original tumor resumed its growth. Finally, by stopping and starting treatment, or by using sub-optimal doses of heparin-cortisone, bladder tumors could be maintained at nearly a constant small size (i.e., 45 to 70 mm 3 ) for periods of more than 8 weeks.
  • a human colon carcinoma was inoculated subcutaneously into nude [athymic] mice, and allowed to grow to a volume of 0.5 cm 3 .
  • Controls and treated animals received the same compounds, except that heparin in the drinking water was 1000 U/ml.
  • an additional treatment group included oral hydrocortisone (0.45 mg/ml) and oral heparin. Animals were housed in cages protected by millipore filter. Tumors grew progressively in all control animals, but regressed in animals treated with heparin and cortisone or heparin and hydrocortisone. The treated tumors were barely palpable after 6 weeks of therapy.
  • CD-1 "Swiss" mice were used because this strain breeds easily and the fertilized females almost always conceive and subsequently deliver a full litter.
  • Groups I, II and IV all produced healthy litters. In Group III there were no mice born and no evidence of abortion. This supports the conclusion that anti-angiogenesis by heparin-cortisone inhibits implantation, presumably by inhibiting capillary growth from the uterus.
  • 17 ⁇ ,2l-Dihydroxypregna-4,9(11)-diene-3,20-dione (or delta-9(11)-deoxycortisol) (90 mg/ml) and Sigma heparin (1.8 mg/ml) were dissolved in 2 cc of 10% ethylene vinyl acetate (EVA) copolymer.
  • EVA ethylene vinyl acetate copolymer
  • a 1-2 mm pellet was cut from the polymer and implanted into the pocket of a rabbit cornea 1 mm from the limbus. The polymer was positioned between the limbal vessels and a 1 mm 3 piece of V2 Carcinoma implanted distal to the pellet. The opposite eye of each of the 3 rabbits tested contained empty polymers.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Molecular Biology (AREA)
  • Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

Angiogenesis in mammals is inhibited by administration of the active agents (1) heparin or a heparin fragment which is a hexasaccharide or larger or analogous compounds and (2) a steroid having 17α- and 21-hydroxy groups, 3- and 20-one groups, and in the 16-position hydrogen hydroxy or a methyl group, and non-toxic physiologically acceptable carboxylates, acetals, ketals and phosphates thereof.

Description

This invention was made with U.S. Government support and the Government has certain rights in the invention.
This application is a divisional of U.S. Ser. No. 07/080,255 filed July 27, 1987, now abandoned which is a continuation of U.S. Ser. No. 06/844,221 filed Mar. 24, 1986, now abandoned which is a continuation of U.S. Ser. No. 06/641,305 filed Aug. 16, 1984 now abandoned which is a continuation-in-part of U.S. Ser. No. 559,175 filed Dec. 7, 1983 which is in turn a continuation-in-part of U.S. Ser. No. 451,431 filed Dec. 20, 1982, now abandoned.
This invention relates to inhibition of angiogenesis and pertains more specifically to treatment of mammals with heparin or heparin fragments or analogues and with steroids to inhibit angiogenesis with subsequent regression of large tumor masses and prevention of tumor metastasis in mammals containing such tumors.
Angiogenesis, the growth of new capillary blood vessels, is important in normal processes such as development of the embryo, formation of the corpus luteum and wound healing. It is also a component in pathologic processes such as chronic inflammation, certain immune responses and neoplasia. Furthermore, angiogenesis is a property of most solid tumors and is necessary for their continued growth.
It has previously been reported that heparin enhanced the intensity of angiogenesis induced by tumors in vivo, although in the absence of tumor cells or tumor extract neither heparin nor the mast cells which release heparin could induce angiogenesis. Taylor and Folkman, Nature Vol. 297, 307-312 (1982). It has also been reported in Shubik et al., J. Natl. Cancer Inst., Vol. 57, 769-774 (1976) that 6 α-methyl-prednisolone partially suppressed tumor angiogenesis in hamster cheek pouch under certain conditions, but tumor growth was not stopped, and many other publications have reported continued growth of tumors even in the presence of large doses of cortisone. It has also been reported in Gross et al., Proc. Natl. Acad. Sci., U.S.A., Vol. 78, 1176-80 (1981) that medroxyprogesterone, dexamethasone, and to a lesser extent cortisone, inhibited tumor angiogenesis in rabbit corneas, while estradiol and testosterone were ineffective
Heparin, an α,β glycosidically linked highly sulfated copolymer of uronic acid and glucosamine, has been used clinically as an anticoagulant for half a century. Despite its importance and widespread use, both the exact structure of heparin and the precise nature by which it acts in blood anticoagulation have not been elucidated. Much of the difficulty in determining the structure of heparin is because it is not a homogeneous substance. Heparin is polydisperse with a molecular weight range from 5,000 to 40,000. Within a given chain, there are also structural variations such as varying degrees of sulfation, N-acetylation, and C-5 epimerization in the uronic acid residue.
Consequently, the precise composition of commercial heparin varies depending on its source and method of purification. Heparin has been degraded by treatment with heparinase (an enzyme of bacterial origin, Langer et al. U.S. Pat. No. 4,341,869) which cleaves the molecule at the α-glycosidic linkages between N-sulfated-D-glucosamine 6-sulfate and L-iduronic acid 2-sulfate to form fragments including disaccharide, tetrasaccharide, hexasaccharide, and larger oligosaccharides, each being simply a chain-shortened heparin fragment with minor end group modification (the degradation results in a Δ-4,5 site of unsaturation in the terminal uronic acid residue). Linhardt et al., J. Biol. Chem., Vol. 257, 7310-13 (1982).
It has now been found that angiogenesis in mammals is inhibited and tumor masses in mammals are caused to regress (and metastasis prevented) by administration of the combination of two essential active agents: (1) heparin or a heparin fragment which is a hexasaccharide or larger, or an analogous compound having one of the structures ##STR1## and (2) a steroid having 17α- and 21-hydroxy groups, 3- and 20-one groups, and in the 16- position hydrogen hydroxy or a methyl group, and non-toxic physiologically acceptable carboxylates, acetals, ketals and phosphates thereof, said combination exhibiting a vascular zone when implanted in an immature chick chorioallantoic membrane.
Among the steroids which are effective are the following:
17α,21-dihydroxy-4-pregnene-3,11,20-trione and its 21-acetate (or cortisone)
11α, 17,21-trihydroxypregn-4-ene-3,20-dione (or 11α-hydrocortisone)
11β,17α,21-trihydroxypregn-4-ene-3,20-dione (or hydrocortisone)
17α,21-dihydroxypregna-4,9(11)-diene-3,20-dione
15α,17α,21-trihydroxy-4-pregnene-3,20-dione
16α,17α,21-trihydroxy-6α -methylpregn-4-ene-3,20-dione-2l-acetate-16,17 cyclic ketal of acetone
6α-fluoro-17α,21-dihydroxy-16β-methylpregna-4,9(11)-diene-3,20-dione
6α-fluoro-17α,21-dihydroxy-16β-methylpregna-4,9(11)-diene-3,20-dione-17,2l-diacetate
6β,17α,21-trihydroxypregn-4-ene-3,20-dione
17α,21-dihydroxypregn-4-ene-3,20-dione-21-acetate
17α,21-dihydroxypregn-4-ene-3,20-dione
9β,11β-epoxy-17α,21-dihydroxy-2α-methylpregn-4-ene-3,20-dione-21-acetate
17α,21-dihydroxy-16α-methylpregn-4-ene-3, 20-dione
9α,11β-dichloro-17α,21-dihydroxypregn-4-ene-3,20-dione-2l-acetate
17α,21-dihydroxy-6α,16α-dimethylpregn-4-ene-3,20-dione-21-acetate
17α,21-dihydroxy-16α-methylpregna-4,9(11)-diene-3,20-dione-2l-acetate
17α,21-dihydroxy-16β-methylpregna-4,9(11)-diene-3,20-dione-2l-benzoate
17α,21-dihydroxy-6β-methylpregna-4,9(11)-diene-3,20-dione-21-acetate
6α-fluoro-17α,21-dihydroxy-16β-methylpregna-4,9(11)-diene-3,20-dione-l7-acetate-21-benzoate
17α,21-dihydroxy-16β-methylpregna-1,4, 9(11)-triene-3,20-dione-17-succinate sodium monohydrate
9α-fluoro-11β,16α,17α,21-tetrahydroxypregn-4-ene-3,20-dione-16,21-diacetate
17α,21-dihydroxy-16α-methylpregna-1,4,9(11)-triene-3,20-dione-21-succinate sodium monohydrate
6α-fluoro-17α,21-dihydroxy-16β-methylpregna-1,4,9(11)-triene-3,20-dione-21-succinate sodium
More preferred are those steroids which lack glucocorticoid and mineralo-corticoid activity, since such activity is an undesired side effect and limits the dose size or extent of use of the steroid for the purpose of the present invention. Among such more preferred steroids are 11α, 17,21-trihydroxypregn-4-ene-3,20-dione (or 11α-hydrocortisone), 17α,21-dihydroxypregn-4-ene-3,20-dione (or 11-desoxycortisol or Cortexolone), and 17α,21-dihydroxypregna-4,9(11)-diene-3,20-dione.
It has also been found that certain steroids which display much greater glucocorticoid activity than hydrocortisone, for example 11β,17α,21-trihydroxy16α-methyl-9α-fluoropregna-1,4-diene-3,20-dione (also known as dexamethasone) and 11β,17α,21-trihydroxypregna-1,4-diene-3,20-dione (also known as prednisolone) are inactive for the purposes of the present invention.
Neither mature non-growing blood vessels nor vascular tissue seems to be affected by the procedure of the present invention. Inhibition of angiogenesis in accordance with the present invention, in addition to its effect upon tumor regression and metastasis in tumor-bearing mammals, is effective as a contraceptive in females even if first administered after insemination has occurred, and is effective in treating diseases involving neovascularization such as neovascular diseases of the eye.
None of the steroids themselves effectively inhibits angiogenesis nor causes regression of tumors in the absence of heparin or a heparin fragment or the specified compounds. Heparin alone does not inhibit angiogenesis but on the contrary enhances it.
The active agents may be mixed together prior to administration or may be administered separately at about the same time so that both are present simultaneously in the mammal being treated. The administration may be oral or parenteral including inter alia topical application, intravenous, intra-arterial or subcutaneous injection, and including absorption as well as injection and introduction into bodily apertures or orifices In the case of heparin, which is commercial available in the form of heparin sodium, oral administration leads to degradation in the gastrointestinal tract which results in loss of its anticoagulant activity, but because it has been found that the degradation products include disaccharide and larger fragments, this mode of administration is highly effective for the present invention both for heparin for heparin fragments, and for the specified compounds. The heparin and its fragments as well as the specified compounds may be employed in any physiologically acceptable non-toxic form, including their metal salts, preferably as the sodium salts, all of which are embraced in the term "heparin" or "fragment" or "compound" as used in the present specification and claims. For best results heparin sold under the trade name "Panheprin" (Abbott Laboratories) is preferred, but heparin from other sources, such as Hepar, Inc. can also be used although less effective
Cortisone and its physiologically acceptable non-toxic derivatives such as the acetate, as well as many other steroids useful in the present invention, are only very slightly soluble in water, hence are preferably administered parenterally, e.g. subcutaneously, not orally. For oral administration, steroids such as hydrocortisone or its 11-α isomer (which are relatively water soluble as compared to cortisone) or one of their water-soluble physiologically acceptable non-toxic derivatives such as a carboxylate, acetal, ketal or the phosphate are preferred Water-insoluble derivatives of water-soluble steroids such as derivatives of hydrocortisone or its 11- α isomer which are non-toxic and physiologically acceptable, are administered parenterally. The terms "cortisone" and "hydrocortisone" and 11- α isomer of hydrocortisone as used in the present specification and claims are intended to include both the steroids themselves and their derivatives as defined above.
Dosages employed are limited only by the well known limits for the administration of the drugs individually for their usual effects, in the case of cortisone, hydrocortisone, or its 11-α isomer. However, a number of the useful steroids have no other biological effect apart from the present invention. Simple testing, for example, by the procedure of
Example 3 below, suffices to determine effectiveness and optimum dose. Heparin may be administered percutaneously in amounts as large as tolerable without objectionable anticoagulant effects. Since heparin administered orally has no anticoagulant effect, and since the hexasaccharide fragment has no anticoagulant effect whether given orally or in any other way, large dosages can thus be administered without risk of bleeding. Oral dosages of heparin of the order of 27,000-45,000 units per kg. body weight per day have been found to be effective, but when administered subcutaneously, doses greater than about 600 units per kg. body weight twice daily led to undesirable anticoagulation effects. In the case of the hexasaccharide fragment, 7 mg per kg. body weight twice daily subcutaneously has been found effective. Cortisone acetate was effective in subcutaneous dosages of 250 mg/kg/day down to 37 mg/kg/day and hydrocortisone was effective orally in amounts of 0.45 mg/ml drinking water (approximately 75 mg/kg/day). The 11- α isomer of hydrocortisone is approximately equal to hydrocortisone in activity for the purpose of the present invention.
The dose size required to bring about regression of tumors or to prevent metastasis varies to some extent depending upon the identity of the tumor, as does the length of time required to bring about complete regression of tumors. Tumor size at the beginning of treatment also affects the length of time required for complete regression. Because of the occurrence of angiogenesis in psoriasis and arthritis, it is expected that the present invention may be useful in treating these diseases. Because administration of cortisone, with or without heparin or heparin fragments or the specified compounds, may result in pulmonary infection after a number of days, it is desirable to administer a suitable antibiotic as prophylaxis during treatment in accordance with the present invention.
The active agents are best dissolved or suspended in a suitable carrier which itself must be non-toxic and physiologically acceptable, such as water or normal saline. Compositions containing mixtures of the active agents, either dry or in a suitable carrier, can be employed.
EXAMPLE 1
Cortisone acetate 0.9 mg in 0.9 ml saline was flooded over the chorioallantoic membrane of 8-day chick embryos through a window in the shell made previously. On day 9, tumor extract (100 μg) from hepatoma cells (as described in Zetter, Nature Vol. 285, 41-43, 1980) in 5 μl H2 O was placed on the center of a round 15 mm diameter plastic coverslip and allowed to dry. To the center of each coverslip was then added a 5 μl aliquot containing either heparin (6 μg, i.e., 1 unit), or water, at least twenty embryos being subjected to each. After drying, the coverslip was placed on the chorioallantoic membrane: Additional control embryos received tumor extract and/or heparin, but were not pre-treated with cortisone acetate. The membranes were viewed on day 11 with a ×12 stereoscope. Angiogenesis was present if new capillaries were seen to converge on the spot in the center of the coverslip. All of the embryos treated with water or heparin but no cortisone exhibited angiogenesis, as well as 80% of those treated with cortisone acetate alone. Less than 2% of those treated with both heparin and cortisone acetate exhibited angiogenesis.
EXAMPLE 2
Porcine mucosal heparin was exhaustively degraded using heparinase by the procedure of Langer et al , Science Vol. 217, 261-3 (1982) and the products were fractionated using Sephadex columns equilibrated with 1M NH4 OAc. The degraded heparin had no anticoagulant activity as determined by activated partial thromboplastic time or whole blood recalcification time. The product or product mixture (250 mg) was dissolved in 1 cc of 1M NH4 OAc, loaded onto a 75×2.5 cm G-15 column, and eluted at 0.5 cc/min. This resulted in several incompletely resolved peaks corresponding to tetra-, hexa-, and higher oligosaccharides and a separate peak corresponding to disaccharide product. The disaccharide peak was freeze-dried, dissolved in 0.2 cc of 1M NH4 OAc and rechromatographed on G-15 resulting in the same sharp peak which was freeze-dried. The mixture of tetra-, hexa-, and oligosaccharides was freeze-dried, redissolved in 1 cc of 1M NH OAc, and eluted from a 50×1.25 cm G-50 at 2 cc/min, resulting in an unresolved double peak corresponding to tetra- and hexasaccharide fragments and an additional peak corresponding to oligosaccharides which was freeze-dried. The tetra- and hexasaccharide fragments were combined, freeze-dried, redissolved in 1 cc of 1M NH4 OAc, and loaded onto a G-15 column. The tetrasaccharide was eluted from a G-15 column in a single peak, the center cut of which was freeze-dried. The hexasaccharide fraction was freeze-dried, redissolved in 0.3 cc 1M NH4 OAc, eluted from a G-15 column in a single peak, the center cut of which was freeze-dried.
Fragment size was determined by dissolving a weighed amount of each fraction into 0.03M hydrochloric acid and measuring the absorbance of this solution at 232 nm. The molecular weight of each fragment was calculated using a molar absorptivity, for the α, β unsaturated carboxylate end group present in each of these products, of ξ=5500. The di- and tetrasaccharides were further characterized by comparing their K avg-values on G-15 with mono-, di-, and trisaccharide standards. Measured molecular weights were 530, 1210, 1600, and 1870 for the di-, tetra-, hexa-, and oligosaccharide fractions respectively.
The various heparin fragments were dissolved into methylcellulose discs either alone or with cortisone acetate. The discs were then applied to the 4-day yolk sac membrane of chick embryos cultured in Petri dishes as described by Taylor and Folkman, Nature, Vol. 297, 307-312 (1982). In the presence of cortisone acetate (100 μg), as shown in the following table, the hexasaccharide fragment demonstrated the highest antiangiogenesis activity.
              TABLE I                                                     
______________________________________                                    
Percent Embryos That Developed Avascular Zones 48 Hours                   
After Implantation of Methylcellulose Discs                               
On The 4-day Old Yolk Sac Membrane                                        
        OLIGO-     HEXA-     TETRA   DI-                                  
CONC.   SAC-       SAC-      SAC-    SAC-                                 
(μg) CHARIDES   CHARIDE   CHARIDE CHARIDE                              
______________________________________                                    
12      all died   100%      0%      0%                                   
8       100%       100%                                                   
4       25%        100%                                                   
1       25%         50%                                                   
0.1       0         50%                                                   
______________________________________
Tetra- and disaccharides were inactive. Oligosaccharides were less active and were toxic at higher concentrations. Therefore, the hexasaccharide fragment was used in subsequent experiments. In the growing 6-day chorioallantoic membrane, discs containing hexasaccharide (12 μg) and cortisone acetate (100 μg) produced large avascular zones up to 12.6±0.1 mm diameter by 48 hours. As in the case of heparin (with cortisone acetate), capillaries in the mesodermal layer were absent while the other two tissue layers of the membrane were intact and viable. Hexasaccharide alone did not promote tumor angiogenesis as heparin did.
All discs contained a combination of cortisone acetate (100 μg) and a heparin fragment. No avascular zones developed in the presence of any heparin fragment alone, or with cortisone or methylcellulose alone. Ten embryos were used for each group. With hexasaccharide (plus cortisone), the area of the avascular zone was 17% of the vascular membrane at 12 μg and 15% at 0.1 μg. For the oligosaccharides, the maximum avascular area was 10%.
EXAMPLE 3
Fertilized chick embryos were removed from their shell on day 3 (or 4) and incubated in a Petri dish in high humidity and 5% CO2 as previously described by Auerbach et al., J. Devel. Biol., Vol. 41, 391-4 (1974), except that an outer dish and antibiotics were not used. On day 6, a methylcellulose disc (10 μl) containing either heparin (6 μg), or hexasaccharide heparin fragment (12 μg), or cortisone acetate (Sigma, powder free of preservatives and suspending agents), or a combination of cortisone acetate+heparin or cortisone acetate+hexasaccharide was implanted on the chorioallantoic membrane. The embryos were examined 48 hours later, and if a clear avascular zone appeared around the methylcellulose disc, the diameter of the zone was measured with a Nikon Profile projector at ×20. Thirty embryos were used in each group. India ink was injected into the heart of some embryos just before formalin fixation so that vessels could be followed to the edge of the avascular zone in histological sections.
Hexasaccharide+cortisone acetate produced avascular zones of 12.6±0.1 mm diameter in all embryos. Heparin+cortisone acetate produced avascular zones of 8.9±0.7 mm diameter. There were no avascular zones in the presence of any compounds alone, or with methylcellulose alone.
Histologic cross-sections of the chorioallantoic membranes, revealed that capillaries developed normally in the presence of any compound alone. In contrast, capillaries were completely absent from the mesodermal layer in the face of either hexasaccharide+cortisone acetate, or heparin+cortisone acetate, while the ectodermal and endodermal cell layers remained unaffected. In the mature chorioallantoic membrane where vessels are no longer growing, the cortisone acetate-heparin or -hexasaccharide fragment combinations were without effect.
Avascular zones were also observed using essentially the same procedure with a combination of heparin (Panheprin) (10 units) with 25-200 μg of each of the following steroids:
11β, 17α, 21-trihydroxypregn-4-ene-3,20-dione
11α, 17α, 21-trihydroxypregn-4-ene-3,20-dione
17α,21-dihydroxypregn-4-ene-3,20-dione
17α,21-dihydroxypregna-4,9(ll)-diene-3,20-dione
9α-fluoro-11α,16α,17α,21-tetrahydroxypregna-1,4-diene-3,20-dione (or triamcinolone)
9α,11β-dichloro-17α,21-dihydroxypregn-4-ene-3,20-dione-21-acetate
17α,21-dihydroxy-16β-methylpregna-4,9(11)-diene-3,20-dione-21-benzoate
17α,21-dihydroxy-16β-methylpregna-1,4,9(11)-triene-3,20-dione-21-succinate sodium monohydrate
6α-fluoro-17α,21-dihydroxy-16β-methylpregna-4,9(11)-diene-3,20-dione
6α-fluoro-17α,21-dihydroxy-16β-methylpregna-4,9(11)-diene-3,20-dione-17,21diacetate
15α,17α,21-trihydroxy-4-pregnene-3,20-dione
16α,17α,21-trihydroxy-6α-methylpregn-4-ene-3,20-dione-21-acetate-16,17-cyclic ketal of acetone
Of the foregoing, the last four displayed the greatest anti-angiogenic activity by this test.
All of the effective combinations of active agents, for the purposes of the present invention, exhibit an avascular zone when implanted on an immature chick chorioallantoic membrane as described in this Example 3.
EXAMPLE 4
Polymer pellets of ethylene vinyl acetate copolymer (EVA) of approximately 1 mm diameter were impregnated, using the procedure of Langer et al., Nature, Vol. 263, 797-800 (1976), with heparin 180 μg (Sigma), or hexasaccharide fragment 300 μg, or cortisone acetate 1.5 mg (Sigma), or a combination of cortisone and heparin. The pellets were implanted in the cornea of a rabbit eye 1 mm from the limbus and a 1 mm3 piece of V2 carcinoma was implanted distal to the polymer, 2 mm from the limbus. In the opposite eye of each rabbit, control pellets that were empty were similarly implanted in juxtaposition to the tumor.
Release rates averaged 15 μg/day for heparin; 21 μg/day for hexasaccharide fragment; and 5 μg/day for cortisone. When the compounds were mixed, they released at the same rates. By spectrophotometry, the pellets released heparin for 14 days, hexasaccharide for 11 days, and cortisone for more than 30 days.
As capillary blood vessels grew towards the tumor implant, maximum vessel length was measured every 3 days with a stereoscopic slit lamp at ×10 (±0.1 mm). On day 14 the rabbits were killed, and India ink was injected into each carotid artery. The corneas were removed and examined with a stereoscope.
New capillary blood vessels were observed growing towards the tumor and passing over an empty pellet or a pellet containing heparin alone at a mean rate of 0.44 mm/day; and over a pellet containing cortisone alone at 0.22 mm/day. The tumors behind these pellets were vascularized by 6-8 days. When the pellets contained both cortisone and heparin, there was no capillary growth for 13 days. When the heparin-cortisone pellets were removed or when the pellets were depleted of heparin, capillary growth resumed. Histologic sections showed that tumor cells remained viable and capable of replication even when they were adjacent to the heparin-cortisone pellet.
In the presence of implanted pellets in which the hexasaccharide fragment of Example 2 replaced the heparin, new capillaries grew toward the tumors at a mean rate of 0.30 mm/day in the presence of the hexasaccharide pellets; 0.14 mm/day when the pellets contained cortisone; and 0.32 mm/day when the pellets were empty. In the presence of the hexasaccharide-cortisone combination, there was no capillary growth throughout the 13-day observation period in 4 rabbits, and in one rabbit a few capillaries grew at 0.07 mm/day.
EXAMPLE 5
(a) Reticulum cell Sarcoma: 1 mm3 pieces of tumor were implanted with a trocar subcutaneously in the backs of 30 mice; 5 per group. Treatment was begun 10 days later, when mean tumor volume was 1.5×102 mm3. Oral heparin was 200 U/ml in drinking water, average daily consumption was 5-10 ml water per 22 g mouse. Cortisone acetate was administered subcutaneously once daily in a dose of 250 mg/kg for six days, 125 mg/kg for one day, 75 mg/kg for one day, and thereafter a daily maintenance dose of 37 mg/kg (a "tapered" dosage). Control animals received either saline injections, or heparin alone or cortisone acetate alone. All controls were dead by day 34 with large primary tumors and lung metastases. All mice treated with oral heparin+cortisone tapered dosage became tumor-free by day 15 and remained so after treatment was discontinued.
An additional group of mice was treated similarly except that heparin was administered twice daily subcutaneously in a dose of 627 units, and cortisone acetate was administered subcutaneously once daily in a uniform dose of 75 mg/kg. Response of the mice was the same as in the first set except that tumors recurred after cessation of treatment; these mice became permanently tumor-free when subjected to the regimen of oral heparin and cortisone acetate tapered dosage described above. One of these mice died on day 31 with no gross primary tumor and no metastases.
(b) Lewis Lung Carcinoma: Treatment began 7 days after implantation of a 1 mm3 piece of tumor in 42 mice: 7 per group. Treatment was with oral heparin and subcutaneous cortisone acetate tapered dosages described above. All controls died by day 33 with large tumors and numerous lung metastases. In the heparin+cortisone acetate groups treatment was discontinued for each mouse after tumor had been invisible for approximately 7 days. In the oral heparin+cortisone acetate all mice were off treatment by day 33, and remained tumor-free. In an additional group treated with subcutaneous heparin and subcutaneous cortisone acetate (75 mg/kg), 5 mice were off treatment at day 37 and remained tumor-free. Two mice died of pneumonia on days 30 and 33 respectively with small primary tumors (<75 mm3). One metastasis was found in one mouse.
To determine if other steroids could substitute for cortisone, heparin was administered with hydrocortisone, dexamethasone, or medroxyprogesterone. Only hydrocortisone was as effective as cortisone acetate in causing tumor regression when administered with heparin. At the highest tolerable doses neither dexamethasone (3.2 mg/Kg), nor medroxyprogesterone (112 mg/Kg), caused regression of Lewis lung tumors with or without heparin.
(c) B-16 Melanoma: 7.4×106 melanoma cells were injected subcutaneously into 40 mice; 5 per group. Treatment of one group was with oral heparin as described above and oral hydrocortisone, 0.45 mg/ml in drinking water Another group received oral heparin and subcutaneous cortisone acetate tapered dosage, and a third group received subcutaneous heparin and cortisone acetate 75 mg/kg. Controls received either water, or heparin alone, or hydrocortisone or cortisone acetate alone. All control animals died by day 31 with large tumors and lung metastases. In the heparin+cortisone acetate groups, treatment was discontinued after tumor had become invisible for approximately 7 days. In the oral heparin+cortisone acetate tapered dosage group, 1 mouse died on day 24 with a partially regressed tumor and 2 lung metastases that were avascular and measured less than 0.1 mm. All other mice in the group became tumor-free and remained so after their treatment was discontinued by day 32. In the subcutaneous heparin+cortisone acetate group one mouse died on day 18 and another on day 21; neither had lung metastases. Treatment was discontinued for the other 3 mice on day 32; tumors 3 weeks later were successfully re-treated with oral heparin+cortisone acetate tapered dosage, and these mice have remained tumor-free In the oral heparin+oral hydrocortisone group, all mice remained tumor-free after their treatment was discontinued on day 47. The regimen of oral heparin+oral hydrocortisone seemed to be more effective for melanoma than it was for ovarian sarcoma or Lewis lung carcinoma.
(d) Bladder Carcinoma: 70 mice: 7 per group received a 1 mm3 implant of tumor subcutaneously. All control animals died by day 31, with large primary tumors. No mice bearing bladder carcinoma developed lung metastases. One group was treated with oral heparin and subcutaneous cortisone acetate tapered dosage, starting on day 9 when mean tumor volume was 140 mm3. Tumors stopped growing, but regressed only partially, and then reached a steady state where tumor volume remained at approximately 70 mm3 for as long as the treatment was continued (i.e., 61 more days). Only one mouse died of pneumonia on day 19. The "dormant" tumors were viable, as evidenced by resumption of tumor growth whenever treatment was discontinued for one mouse at a time, beginning at day 70. For a second group treated with subcutaneous heparin and subcutaneous cortisone acetate 75 mg/kg as described above, the result was similar; i.e., long-term tumor "dormancy". One mouse died at day 21.
Because of the inability the standard regimen of oral heparin+cortisone acetate to bring about complete regression, higher concentrations of oral heparin were used with other groups. With oral heparin (600 U/ml)-cortisone acetate tapered dosage, there was more significant tumor regression and a steady state ("dormancy") was reached at a smaller tumor volume of approximately 45 mm3. One mouse died. However, with 1000 U/ml heparin, there was complete regression; mice remained tumor-free after discontinuation of treatment on ay 39. No mice died in this group.
In summary, all tumors either stopped growing or regressed when the heparin-cortisone acetate combination was administered. In contrast, when either compound was used alone, tumor growth continued at the same rate as in animals receiving only saline injections; all such control animals died with a large tumor burden.
In the majority of animals treated with heparin+cortisone acetate, it was possible to achieve "complete regression", i.e., tumors did not recur after treatment was discontinued. Thus, with the most effective regimen, oral heparin (200 U/ml)+cortisone (s.c. 250-tapered dosage), it was possible to obtain "complete regression" in 100% of ovarian sarcomas, 100% of Lewis lung carcinomas and 80% of B-16 melanomas. However, when bladder carcinomas were treated with this regimen, there were no "complete regressions" until heparin was increased to 1000 U/ml, following which 100% of tumors regressed without recurrence. With the less effective regimen, heparin (s.c.)+cortisone (75 mgm), complete regression rate was ovarian, 80%; Lewis lung, 71%; B-16 melanoma 60%; and bladder 0%.
EXAMPLE 6
The hexasaccharide heparin fragment of Example 2 was dissolved in saline, 1.5 mg/ml. Three mice bearing implanted ovarian sarcoma were treated with subcutaneous injection of the hexasaccharide fragment twice daily at a dosage of 7 mg/kg and subcutaneous injection of cortisone acetate- tapered dosage. Control mice received either cortisone acetate alone or saline. While control tumors grew progressively, the hexasaccharide+cortisone acetate treated tumors regressed rapidly and were barely visible 4 days later. Their treatment with hexasaccharide was then discontinued, and the tumors reappeared 3-5 days later.
EXAMPLE 7
To directly observe avascular tumors during systemic therapy, Lewis lung tumors were implanted in the mouse cornea by the procedure of Muthukkaruppan et al., Science, Vol. 205, 1416-18 (1979) and treatment was begun 24 hours later. Heparin (oral)-cortisone acetate tapered dosage significantly inhibited capillary growth (0.02 mm/day) compared to cortisone acetate alone (0.24 mm/day), heparin alone (0.32 mm/day) or saline (0.23 mm/day). In the presence of heparin-cortisone acetate, a thin plate of tumor remained avascular. Three-dimensional tumor-growth did not occur. In contrast, in the saline controls or when either heparin or cortisone acetate were administered alone, tumors became vascularized and grew as a three-dimensional mass until they eventually perforated the cornea. These large tumors could be regressed to the flat, thin intracorneal phase by the resumption of the heparin-cortisone acetate combination. However, the intracorneal tumor cells could not be eradicated; discontinuation of the heparin-cortisone acetate led to recurrence of a vascularized tumor.
Lung metastases were counted in all animals that died. A ×6 stereoscope was used. In all control animals, the lungs were heavily studded with metastases from the three types of metastasizing tumors. In contrast, when any combination of heparin+cortisone acetate was used, no metastases were found in mice bearing ovarian sarcoma; only 1 metastasis was found in a mouse bearing Lewis lung carcinoma; and 2 avascular metastases less than 0.1 mm diameter were found in one mouse bearing B-16 melanoma. The nearly complete absence of metastases in heparin+cortisone treated mice was so striking, that the effect can be more readily appreciated by the following expression of data
Total Number Lung Metastases 
______________________________________                                    
 Total Number Lung Metastases                                             
______________________________________                                    
Controls =            4553 in 73 animals                                  
Heparin + Cortisone =                                                     
                     3 in 39 animals                                      
______________________________________
Furthermore, no lung metastases appeared in any surviving animals that were off treatment.
To exclude the possibility that tumor regression might be caused by direct cytotoxicity, all 4 types of tumor cells were cultured in the presence of 10% serum obtained from mice receiving either heparin, cortisone acetate, heparin-cortisone acetate, or no drug. Heparin-cortisone acetate did not inhibit cell growth, but in fact stimulated it. Furthermore, histological sections shows no evidence of a cytotoxic effect on bone marrow or intestinal mucosa in animals receiving heparin-cortisone acetate.
To exclude the possibility that heparin-cortisone acetate might induce tumor regression by promoting an immune reaction, mice were inoculated with fresh tumor cells at various intervals after they were off treatment. These tumors grew at the same rate as the original implants. Furthermore, if tumor regression was nearly complete and heparin-cortisone acetate was discontinued prematurely, the original tumor resumed its growth. Finally, by stopping and starting treatment, or by using sub-optimal doses of heparin-cortisone, bladder tumors could be maintained at nearly a constant small size (i.e., 45 to 70 mm3) for periods of more than 8 weeks.
Inflammatory angiogenesis induced by implantation of silica particles into the rabbit cornea, immune angiogenesis induced by implantation of lymph node from a different rabbit, were also completely prevented by the cortisone-heparin pellets. Cortisone by itself temporarily delayed the onset of both types of angiogenesis (compared to an empty pellet), and heparin by itself delayed the onset of immune angiogenesis, but neither alone prevented angiogenesis for an extended period of time as did the cortisone-heparin combination.
EXAMPLE 8
A human colon carcinoma was inoculated subcutaneously into nude [athymic] mice, and allowed to grow to a volume of 0.5 cm3. Controls and treated animals received the same compounds, except that heparin in the drinking water was 1000 U/ml. also an additional treatment group included oral hydrocortisone (0.45 mg/ml) and oral heparin. Animals were housed in cages protected by millipore filter. Tumors grew progressively in all control animals, but regressed in animals treated with heparin and cortisone or heparin and hydrocortisone. The treated tumors were barely palpable after 6 weeks of therapy.
EXAMPLE 9
CD-1 "Swiss" mice were used because this strain breeds easily and the fertilized females almost always conceive and subsequently deliver a full litter.
One male was left alone in the cage for 24 hours. His bedding was then changed and two females were added at 5 p.m. The females were then checked at 8 a.m. the next morning for the presence of a cervical plug, which indicates insemination. Sufficient male-female cages were set up to obtain at least 20 pregnant females.
Treatment of the inseminated females was started on the day after insemination by offering drinking water to each of four groups, as follows:
______________________________________                                    
Group I        Heparin (Hepar) 1,000 U/ml                                 
               in water                                                   
II             Hydrocortisone phosphate,                                  
               0.45 mg/ml in water                                        
III            (I) and (II) together in water                             
IV             Water alone                                                
______________________________________
The treatments were continued for only four days, after which female mice were placed one per cage, and followed closely for the presence of offspring. The cages were checked daily for any sign of abortion (fur, fetal remains, etc.).
Groups I, II and IV all produced healthy litters. In Group III there were no mice born and no evidence of abortion. This supports the conclusion that anti-angiogenesis by heparin-cortisone inhibits implantation, presumably by inhibiting capillary growth from the uterus.
EXAMPLE 10 Inhibition of Tumor Vessels in the Rabbit Cornea
17α,2l-Dihydroxypregna-4,9(11)-diene-3,20-dione (or delta-9(11)-deoxycortisol) (90 mg/ml) and Sigma heparin (1.8 mg/ml) were dissolved in 2 cc of 10% ethylene vinyl acetate (EVA) copolymer. A 1-2 mm pellet was cut from the polymer and implanted into the pocket of a rabbit cornea 1 mm from the limbus. The polymer was positioned between the limbal vessels and a 1 mm3 piece of V2 Carcinoma implanted distal to the pellet. The opposite eye of each of the 3 rabbits tested contained empty polymers. Mean vessel growth or 0.45 mm/day over a period of 16 days was seen with the control corneas and 0.07 mm/day was seen in the eyes containing steroid and heparin Measurements are expressed as the mean±SD. Similar results have been demonstrated with cortexolone and heparin implanted in the same manner.

Claims (15)

What is claimed is:
1. A composition for treating mammals for stopping growth, bringing about regression, or preventing metastasis of solid tumors in mammals by oral, or parenteral administration in which the active agents consist essentially of effective proportions for said treating of (1) a member selected from the group consisting of a
said composition including a non-toxic physiologically acceptable carrier adapted for oral or parenteral administration,
said composition exhibiting an avascular zone when implanted in an immature chick chorioallantoic membrane. heparin fragment which is either a hexasaccharide or larger oligosaccharide and an analogous compound having of the structures ##STR2## and (2) a member selected from the group consisting of steroids having a 17α- and 21-hydroxy groups, 3- and 20-one groups, and in the 16-position hydrogen, hydroxy, or a methyl group, and a non-toxic physiologically acceptable carboxylate, or acetal, or ketal, or phosphate thereof.
2. A composition as claimed in claim 1 in which the active agents consist essentially of a heparin fragment which is either a hexasaccharide or larger oligosaccharide and cortisone.
3. A composition as claimed in claim 1 in which the active agents consist essentially of a heparin fragment which is either hexasaccharide or larger oligosaccharide and hydrocortisone.
4. A composition for treating mammals for stopping growth, bringing about regression, or preventing metastasis of solid tumors in mammals by oral administration only, in which the active agents consist essentially of effective proportions for said treating of (1) a member selected from the group consisting of heparin and a heparin fragment which is either a hexasaccharide or larger oligosaccharide and an analogous compound having one of the structures ##STR3## and (2) a member selected from the group consisting of steroids having 17α- and 21-hydroxy groups, 3- and in the 16-position hydrogen, hydroxy, or a methyl group, and a non-toxic physiologically acceptable carboxylate, or acetal, or ketal, or phosphate thereof, said composition including a non-toxic physiologically acceptable carrier adapted for oral administration only. said composition exhibiting an avascular zone when implanted in an immature chick chorioallantoic membrane.
5. The composition as claimed in claim 4 in which the active agents consist essentially of (1) heparin and (2) cortisone.
6. The composition as claimed in claim 4 in which the active agents consist essentially of (1) heparin and (2) hydrocortisone.
7. A method of treating mammals bearing solid tumor masses to stop growth, to bring about regression or to prevent metastasis thereof which comprises administering orally thereto a dose effective for said treating of active agents consisting essentially of (1) heparin or a heparin fragment which is a hexasaccharide or larger oligosaccharide or an analogous compound having one of the structures ##STR4## and (2) a steroid having 17 α- and 21-hydroxy groups, 3- and 20-one groups, and in the 16- position hydrogen, hydroxy, or a methyl group, and a non-toxic physiologically acceptable carboxylate, or acetal, or ketal, or phosphate thereof, wherein said tumor masses are susceptible to said treatment with said active agents.
8. The method as claimed in claim 7 in which administration is oral and the active agents consists essentially of heparin and hydrocortisone.
9. The method as claimed in claim 7 in which administration is oral and the active agents consist essentially of a hexasaccharide heparin fragment and hydrocortisone.
10. The method as claimed in claim 7 in which said administration is oral and said active agent (1) is heparin.
11. A method of treating mammals bearing solid tumor masses to stop growth, to bring about regression or to prevent metastasis thereof which comprises administering thereto an effective dose effective for said treating of active agents consisting essentially of (1) a heparin fragment which is either a hexasaccharide or larger oligosaccharide or an analogous compound having one of the structures ##STR5## and (2) a steroid having 17α- and 21-hydroxy groups, 3- and 20-one groups, and in the 16- position hydrogen, hydroxy, or a methyl group, and a non-toxic physiologically acceptable carboxylate, or acetal, or ketal, or phosphate thereof, wherein said tumor masses are susceptible to said treatment with said active agents.
12. The method as claimed in claim 11 in which the active agents consist essentially of (1) either said heparin fragment or analogous compound and (2) cortisone.
13. The method as claimed in claim 11 in which the active agents consist essentially of (1) said heparin fragment and (2) cortisone.
14. The method as claimed in claim 11 in which the active agents consists essentially of (1) either said heparin fragment or analogous compound and (2) hydrocortisone.
15. The method as claimed in claim 11 in which said active agents consist essentially of (1) said heparin fragment and (2) hydrocortisone.
US07/257,409 1982-12-20 1988-10-13 Inhibition of angiogenesis Expired - Fee Related US4994443A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
US07/257,409 US4994443A (en) 1982-12-20 1988-10-13 Inhibition of angiogenesis

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US45143182A 1982-12-20 1982-12-20
US55917583A 1983-12-07 1983-12-07
US8025587A 1987-07-27 1987-07-27
US07/257,409 US4994443A (en) 1982-12-20 1988-10-13 Inhibition of angiogenesis

Related Parent Applications (1)

Application Number Title Priority Date Filing Date
US8025587A Division 1982-12-20 1987-07-27

Publications (1)

Publication Number Publication Date
US4994443A true US4994443A (en) 1991-02-19

Family

ID=27491531

Family Applications (1)

Application Number Title Priority Date Filing Date
US07/257,409 Expired - Fee Related US4994443A (en) 1982-12-20 1988-10-13 Inhibition of angiogenesis

Country Status (1)

Country Link
US (1) US4994443A (en)

Cited By (91)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5474765A (en) * 1992-03-23 1995-12-12 Ut Sw Medical Ctr At Dallas Preparation and use of steroid-polyanionic polymer-based conjugates targeted to vascular endothelial cells
US5593990A (en) * 1993-03-01 1997-01-14 The Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis
US5721226A (en) * 1993-03-10 1998-02-24 Magainin Pharmaceuticals Inc. Method for inhibiting angiogenesis using squalamine and squalamine steroid derivatives
US5763430A (en) * 1995-06-07 1998-06-09 Magainin Pharmaceuticals Inc. Method of treating a viral infection by administering a steroid compound
US5792635A (en) * 1995-06-07 1998-08-11 Magainin Pharmaceuticals, Inc. Method of inhibiting the sodium/proton exchanger NHE3 and method of inhibiting growth by administering squalamine
US5795885A (en) * 1995-06-07 1998-08-18 Magainin Pharmaceuticals Inc. Method of inhibiting profileration of cells by administering an aminosterol compound
US5840740A (en) * 1995-06-07 1998-11-24 Magainin Pharmaceuticals Inc. Aminosterol compounds and a method of treating infection using the aminosterol compounds
US5840936A (en) * 1995-06-07 1998-11-24 Magainin Pharmaceuticals Inc. Aminosterol compounds useful as inhibitors of the sodium/proton exchanger (NHE)
US5847172A (en) * 1995-06-07 1998-12-08 Magainin Pharmaceuticals Inc. Certain aminosterol compounds and pharmaceutical compositions including these compounds
US5853746A (en) * 1991-01-31 1998-12-29 Robert Francis Shaw Methods and compositions for the treatment and repair of defects or lesions in cartilage or bone using functional barrier
US5856535A (en) * 1994-08-18 1999-01-05 Magainin Pharmaceuticals, Inc. Aminosterol ester compounds
US5874597A (en) * 1995-06-07 1999-02-23 Magainin Pharmaceuticals, Inc. Certain aminosterol compounds and pharmaceutical compositions including these compounds
US5981471A (en) * 1997-02-06 1999-11-09 Entremed, Inc. Compositions and methods for inhibiting cellular proliferation
US5994336A (en) * 1995-06-07 1999-11-30 Magainin Pharmaceuticals Inc. Method of inhibiting proliferation of cells by administering an aminosterol compound
US6090794A (en) * 1990-04-19 2000-07-18 The General Hospital Corporation Inhibition of neurofibrosarcoma growth and angiogenesis
US6114355A (en) * 1993-03-01 2000-09-05 D'amato; Robert Methods and compositions for inhibition of angiogenesis
US6143738A (en) * 1995-06-07 2000-11-07 Magainin Pharmaceuticals, Inc. Therapeutic uses for an aminosterol compound
US6147060A (en) * 1996-04-26 2000-11-14 Magainin Pharmaceuticals Treatment of carcinomas using squalamine in combination with other anti-cancer agents
US6150407A (en) * 1998-03-25 2000-11-21 Large Scale Biology Corporation Methods for inhibiting angiogenesis
US6228879B1 (en) 1997-10-16 2001-05-08 The Children's Medical Center Methods and compositions for inhibition of angiogenesis
US6262283B1 (en) 1996-12-06 2001-07-17 Magainin Pharmaceuticals Inc. Stereoselective synthesis of 24-hydroxylated compounds useful for the preparation of aminosterols, vitamin D analogs, and other compounds
US20020002294A1 (en) * 1997-09-24 2002-01-03 D' Amato Robert J. Estrogenic compounds as antiangiogenic agents
US6420345B1 (en) 1999-03-01 2002-07-16 Cell Genesys, Inc. Methods and reagents for inhibiting angiogenesis
US6433012B1 (en) * 1998-03-25 2002-08-13 Large Scale Biology Corp. Method for inhibiting inflammatory disease
US6506411B2 (en) 1993-07-19 2003-01-14 Angiotech Pharmaceuticals, Inc. Anti-angiogenic compositions and methods of use
US20030096800A1 (en) * 1993-08-06 2003-05-22 D'amato Robert John Estrogenic compounds as anti-mitotic agents
US6596712B2 (en) 1996-04-26 2003-07-22 Genaera Corporation Treatment of carcinomas using squalamine in combination with other anti-cancer agents or modalities
US6605622B2 (en) 1998-08-11 2003-08-12 Entremed, Inc. Use of anti-estrogenic compounds as anti-fungal agents
US20030164219A1 (en) * 2002-02-20 2003-09-04 Joerg Brahm Headliner/duct assembly and welding process therefor
US20030171576A1 (en) * 2000-04-12 2003-09-11 Kinney William A. Regioselective and stereoselective oxidation of fused ring systems useful for the preparation of aminosterols
US20030191098A1 (en) * 1996-11-05 2003-10-09 D'amato Robert J. Methods and compositions for inhibition of angiogenesis
WO2003097052A2 (en) 2002-05-17 2003-11-27 Celgene Corporation Methods and compositions using immunomodulatory compounds for treatment and management of cancers and other diseases
US20030236439A1 (en) * 2002-01-30 2003-12-25 Gregory Agoston Non-steroidal analogs of 2-methoxyestradiol
US6673828B1 (en) 1998-05-11 2004-01-06 Children's Medical Center Corporation Analogs of 2-Phthalimidinoglutaric acid
US20040029955A1 (en) * 2002-05-15 2004-02-12 Takashi Kouge Composition for use in prevention or treatment of vascular-related diseases
US20040037875A1 (en) * 2000-07-12 2004-02-26 Metselaar Josbert Maarten Composition for treatment of inflammatory disorders
US6723858B2 (en) 1993-08-06 2004-04-20 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US20040097479A1 (en) * 1999-08-23 2004-05-20 Agoston Gregory E. Methods of obtaining 2-methoxyestradiol of high purity
US20040147558A1 (en) * 2000-11-30 2004-07-29 Anthony Treston Synthesis of 3-amino-thalidomide and its enantiomers
US20040214807A1 (en) * 1993-08-06 2004-10-28 D'amato Robert J. Estrogenic compounds as anti-mitotic agents
US20050014737A1 (en) * 2003-05-28 2005-01-20 Agoston Gregory E. Antiangiogenic agents
US20050192258A1 (en) * 2000-08-18 2005-09-01 Agoston Gregory E. Antiangiogenic agents
US20050203075A1 (en) * 2004-03-12 2005-09-15 Agoston Gregory E. Antiangiogenic agents
US20050202078A1 (en) * 2002-08-27 2005-09-15 Schiffelers Raymond M. Vesicle-encapsulated corticosteroids for the treatment of cancer
US6946439B2 (en) 1997-02-06 2005-09-20 Entre Med, Inc. Compositions and methods for inhibiting cellular proliferation comprising TFPI fragments
US20050282253A1 (en) * 1995-10-23 2005-12-22 Folkman M J Therapeutic antiangiogenic endostatin compositions
US20060025329A1 (en) * 1997-02-06 2006-02-02 Todd Hembrough Compositions and methods for inhibiting cellular proliferation comprising TFPI fragments
US6995278B2 (en) 2000-08-18 2006-02-07 Entre Med, Inc. Antiangiogenic agents
US20060116360A1 (en) * 2004-11-29 2006-06-01 Fogler William E Method of administering anti-angiogenic agents and a method of treating disease using same
US7135581B2 (en) 2000-08-18 2006-11-14 Entremed, Inc. Antiangiogenic agents
US20070004689A1 (en) * 2004-03-12 2007-01-04 Agoston Gregory E Antiangiogenic agents
US20070020271A1 (en) * 2003-03-04 2007-01-25 Beverly Teicher Endothelial cell specific antibodies and uses thereof
WO2007027527A2 (en) 2005-08-31 2007-03-08 Celgene Corporation Isoindole-imide compounds and compositions comprising and methods of using the same
US20070111972A1 (en) * 2005-07-12 2007-05-17 Dmi Biosciences, Inc. "methods and products for treatment of diseases"
US20070176403A1 (en) * 2006-02-01 2007-08-02 Dennis Calderone Air adjustable seat
US20070185069A1 (en) * 2005-11-14 2007-08-09 Plum Stacy M Anti-angiogenic activity of 2-methoxyestradiol in combination with anti-cancer agents
US20070203066A1 (en) * 2005-12-09 2007-08-30 Holmes Christopher P Compositions and methods for inhibiting cellular proliferation
US20070231410A1 (en) * 2006-03-20 2007-10-04 Fogler William E Disease modifying anti-arthritic activity of 2-Methoxyestradiol
US20080234243A1 (en) * 2007-01-31 2008-09-25 Lavallee Theresa M Method of treating amyloidosis mediated diseases
WO2009042177A1 (en) 2007-09-26 2009-04-02 Celgene Corporation 6-, 7-, or 8-substituted quinazolinone derivatives and compositions comprising and methods of using the same
EP2077112A1 (en) 2003-11-06 2009-07-08 Celgene Corporation Methods and compositions using thalidomide for the treatment and management of cancers and idiopathic pulmonary fibrosis
US20090226509A1 (en) * 2002-06-12 2009-09-10 Enceladus Pharmaceuticals B.V. Composition for treatment of inflammatory disorders
US20090298882A1 (en) * 2008-05-13 2009-12-03 Muller George W Thioxoisoindoline compounds and compositions comprising and methods of using the same
US20100087408A1 (en) * 2008-05-28 2010-04-08 Validus Genetics NON-HORMONAL STEROID MODULATORS OF NF-kB FOR TREATMENT OF DISEASE
WO2010093434A1 (en) 2009-02-11 2010-08-19 Celgene Corporation Isotopologues of lenalidomide
EP2258363A1 (en) 2002-05-17 2010-12-08 Celgene Corporation Compositions for treatment of cancers
US20100323991A1 (en) * 2009-06-22 2010-12-23 Dmi Acquisition Corp. Methods and products for treatment of diseases
US20100324005A1 (en) * 2009-06-22 2010-12-23 Dmi Acquisition Corp. Method for treatment of diseases
WO2011100380A1 (en) 2010-02-11 2011-08-18 Celgene Corporation Arylmethoxy isoindoline derivatives and compositions comprising and methods of using the same
EP2420498A1 (en) 2006-09-26 2012-02-22 Celgene Corporation 5-substituted quinazolinone derivatives as anti-cancer agents
US8143283B1 (en) 1993-03-01 2012-03-27 The Children's Medical Center Corporation Methods for treating blood-born tumors with thalidomide
EP2460522A2 (en) 2003-05-15 2012-06-06 Celgene Corporation Methods and compositions using immunomodulatory compounds for treatment and management of cancers and other diseases
WO2012125438A1 (en) 2011-03-11 2012-09-20 Celgene Corporation Solid forms of 3-(5-amino-2methyl-4-oxo-4h-quinazolin-3-yl)-piperidine-2,6-dione, and their pharmaceutical compositions and uses
WO2012149299A2 (en) 2011-04-29 2012-11-01 Celgene Corporaiton Methods for the treatment of cancer and inflammatory diseases using cereblon as a predictor
WO2012177678A2 (en) 2011-06-22 2012-12-27 Celgene Corporation Isotopologues of pomalidomide
EP2749559A1 (en) 2008-05-30 2014-07-02 Celgene Corporation 5-substituted isoindoline compounds
US9198921B2 (en) 2010-04-05 2015-12-01 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-κB for treatment of disease
EP2985281A2 (en) 2008-10-29 2016-02-17 Celgene Corporation Isoindoline compounds for use in the treatment of cancer
US9351979B2 (en) 2012-12-19 2016-05-31 Ampio Pharmaceuticals, Inc. Methods of treatment of diseases
EP3101017A1 (en) 2007-03-20 2016-12-07 Celgene Corporation 4'-o-substituted isoindoline derivatives and compositions comprising and methods of using the same
US9587281B2 (en) 2012-08-14 2017-03-07 Celgene Corporation Cereblon isoforms and their use as biomarkers for therapeutic treatment
US9611465B2 (en) 2009-05-25 2017-04-04 Celgene Corporation Pharmaceutical composition containing core factor involved in proliferation and differentiation of central nervous cell
WO2017120415A1 (en) 2016-01-08 2017-07-13 Celgene Corporation Solid forms of 2-(4-chlorophenyl)-n-((2-2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl) methyl)-2,2-difluoroacetamide, and their pharmaceutical compositions and uses
US9857359B2 (en) 2012-06-29 2018-01-02 Celgene Corporation Methods for determining drug efficacy using cereblon-associated proteins
US9938254B2 (en) 2016-01-08 2018-04-10 Celgene Corporation Antiproliferative compounds, and their pharmaceutical compositions and uses
US10034872B2 (en) 2014-08-22 2018-07-31 Celgene Corporation Methods of treating multiple myeloma with immunomodulatory compounds in combination with antibodies
US10092555B2 (en) 2014-06-27 2018-10-09 Celgene Corporation Compositions and methods for inducing conformational changes in cereblon and other E3 ubiquitin ligases
EP3524598A1 (en) 2012-08-09 2019-08-14 Celgene Corporation A solid form of (s)-3-(4-((4-morpholinomethyl)benzyl)oxy)-1-oxoisoindolin-2-yl)piperidine-2,6-dione hydrochloride
US10799514B2 (en) 2015-06-29 2020-10-13 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-kappa beta for treatment of disease
US10830762B2 (en) 2015-12-28 2020-11-10 Celgene Corporation Compositions and methods for inducing conformational changes in cereblon and other E3 ubiquitin ligases
US11382922B2 (en) 2019-03-07 2022-07-12 Reveragen Biopharma, Inc. Aqueous oral pharmaceutical suspension compositions

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4771042A (en) * 1985-11-25 1988-09-13 The Upjohn Company Inhibition of angiogenesis involving the coadministration of steroids with heparin or heparin fragments

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4771042A (en) * 1985-11-25 1988-09-13 The Upjohn Company Inhibition of angiogenesis involving the coadministration of steroids with heparin or heparin fragments

Non-Patent Citations (10)

* Cited by examiner, † Cited by third party
Title
Eichbaum Rev. Bras. de Pesquisas Med. Biol. 8(5 6): 489 496, 1975. *
Eichbaum Rev. Bras. de Pesquisas Med. Biol. 8(5-6): 489-496, 1975.
Elias et al., Cancer Chemotherapy Reports, Part 1, vol. 56, No. 6, 12/72, pp. 783 785. *
Elias et al., Cancer Chemotherapy Reports, Part 1, vol. 56, No. 6, 12/72, pp. 783-785.
Elias et al., Cancer, 7/75, vol. 36, pp. 129 136. *
Elias et al., Cancer, 7/75, vol. 36, pp. 129-136.
Grossman et al., Proc. Natl. Acad. Sci. U.S.A., vol. 78, 1176 1180 (1981). *
Grossman et al., Proc. Natl. Acad. Sci. U.S.A., vol. 78, 1176-1180 (1981).
The Merck Index, 9th edition, 1976, pp. 629 630 & 999. *
The Merck Index, 9th edition, 1976, pp. 629-630 & 999.

Cited By (216)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6090794A (en) * 1990-04-19 2000-07-18 The General Hospital Corporation Inhibition of neurofibrosarcoma growth and angiogenesis
US6362392B1 (en) 1990-04-19 2002-03-26 The General Hospital Corporation Nude mouse model for the growth and treatment of human neurally-derived tumors
US5853746A (en) * 1991-01-31 1998-12-29 Robert Francis Shaw Methods and compositions for the treatment and repair of defects or lesions in cartilage or bone using functional barrier
US5762918A (en) * 1992-03-23 1998-06-09 Board Of Regents The University Of Texas System Methods of using steroid-polyanionic polymer-based conjugated targeted to vascular endothelial cells
US5474765A (en) * 1992-03-23 1995-12-12 Ut Sw Medical Ctr At Dallas Preparation and use of steroid-polyanionic polymer-based conjugates targeted to vascular endothelial cells
EP1264597A2 (en) 1993-03-01 2002-12-11 The Children's Medical Center Corporation Methods and compositions for treating cancer
US5629327A (en) * 1993-03-01 1997-05-13 Childrens Hospital Medical Center Corp. Methods and compositions for inhibition of angiogenesis
US20050004087A1 (en) * 1993-03-01 2005-01-06 The Children's Medical Center Corporation Methods and compositions for treating an ocular neovascular disease
US8012996B2 (en) 1993-03-01 2011-09-06 Children's Medical Center Corporation Methods and composition for inhibition of angiogenesis
US20070021464A1 (en) * 1993-03-01 2007-01-25 D Amato Robert Methods and compositions for inhibition of angiogenesis with EM-12 Derivatives
US20030176463A1 (en) * 1993-03-01 2003-09-18 D'amato Robert Methods and compositions for inhibition of angiogenesis with phthaloyl glutamic acid derivatives
US7723361B2 (en) 1993-03-01 2010-05-25 Celgene Corporation Methods for inhibiting undesired angiogenesis in patients having tumors with thalidomide
US20020061923A1 (en) * 1993-03-01 2002-05-23 D'amato Robert Methods and compositions for inhibition of angiogenesis with EM-138
US6235756B1 (en) 1993-03-01 2001-05-22 The Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis by thalidomide
US5712291A (en) * 1993-03-01 1998-01-27 The Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis
US6469045B1 (en) 1993-03-01 2002-10-22 The Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis with EM-138
US8143283B1 (en) 1993-03-01 2012-03-27 The Children's Medical Center Corporation Methods for treating blood-born tumors with thalidomide
US6977268B2 (en) 1993-03-01 2005-12-20 Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis with EM-138
US6071948A (en) * 1993-03-01 2000-06-06 The Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis
US5593990A (en) * 1993-03-01 1997-01-14 The Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis
US6114355A (en) * 1993-03-01 2000-09-05 D'amato; Robert Methods and compositions for inhibition of angiogenesis
US20050215593A1 (en) * 1993-03-01 2005-09-29 D Amato Robert Methods treating tumors with thalidomide
USRE40360E1 (en) * 1993-03-01 2008-06-03 The Children's Medical Center Corporation Methods for inhibition of angiogenesis with a composition comprising 3-hydroxythalidomide
US7524865B2 (en) 1993-03-01 2009-04-28 Celgene Corporation Methods and compositions for treating an ocular neovascular disease
US6420414B1 (en) 1993-03-01 2002-07-16 The Children's Medical Center Corporation Amino derivatives of EM-138 and methods of treating angiogenesis with same
US5721226A (en) * 1993-03-10 1998-02-24 Magainin Pharmaceuticals Inc. Method for inhibiting angiogenesis using squalamine and squalamine steroid derivatives
US5733899A (en) * 1993-03-10 1998-03-31 Magainin Pharmaceuticals Inc. Method for treating infection using steroid based pharmaceutical compositions
US6506411B2 (en) 1993-07-19 2003-01-14 Angiotech Pharmaceuticals, Inc. Anti-angiogenic compositions and methods of use
US7081477B2 (en) 1993-08-06 2006-07-25 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US20060079576A1 (en) * 1993-08-06 2006-04-13 D Amato Robert J Estrogenic compounds as anti-mitotic agents
US20040214807A1 (en) * 1993-08-06 2004-10-28 D'amato Robert J. Estrogenic compounds as anti-mitotic agents
US6930128B2 (en) 1993-08-06 2005-08-16 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US7381848B2 (en) 1993-08-06 2008-06-03 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US7012070B2 (en) 1993-08-06 2006-03-14 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US7109187B2 (en) 1993-08-06 2006-09-19 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US20050020555A1 (en) * 1993-08-06 2005-01-27 D'amato Robert John Estrogenic compounds as anti-mitotic agents
US6723858B2 (en) 1993-08-06 2004-04-20 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US20030096800A1 (en) * 1993-08-06 2003-05-22 D'amato Robert John Estrogenic compounds as anti-mitotic agents
US7291610B2 (en) 1993-08-06 2007-11-06 The Children's Medical Center Corporation Estrogenic compounds as anti-mitotic agents
US20030236408A1 (en) * 1993-08-06 2003-12-25 D'amato Robert John Estrogenic compounds as anti-mitotic agents
US20060183727A1 (en) * 1993-08-06 2006-08-17 D Amato Robert J Estrogenic compounds as anti-mitotic agents
US5856535A (en) * 1994-08-18 1999-01-05 Magainin Pharmaceuticals, Inc. Aminosterol ester compounds
US6143738A (en) * 1995-06-07 2000-11-07 Magainin Pharmaceuticals, Inc. Therapeutic uses for an aminosterol compound
US5795885A (en) * 1995-06-07 1998-08-18 Magainin Pharmaceuticals Inc. Method of inhibiting profileration of cells by administering an aminosterol compound
US5994336A (en) * 1995-06-07 1999-11-30 Magainin Pharmaceuticals Inc. Method of inhibiting proliferation of cells by administering an aminosterol compound
US20050261508A1 (en) * 1995-06-07 2005-11-24 Magainin Pharmaceuticals Inc. Aminosterol compounds useful as inhibitors of the sodium/proton exchanger (NHE), pharmaceutical methods and compositions employing such inhibitors, and processes for evaluating the NHE-inhibitory efficacy of compounds
US5763430A (en) * 1995-06-07 1998-06-09 Magainin Pharmaceuticals Inc. Method of treating a viral infection by administering a steroid compound
US5840936A (en) * 1995-06-07 1998-11-24 Magainin Pharmaceuticals Inc. Aminosterol compounds useful as inhibitors of the sodium/proton exchanger (NHE)
US20060183928A1 (en) * 1995-06-07 2006-08-17 Genaera Corporation Aminosterol compounds useful as inhibitors of the sodium/proton exchanger (NHE), pharmaceutical methods and compositions employing such inhibitors, and processes for evaluating the NHE-inhibitory efficacy of compounds
US5847172A (en) * 1995-06-07 1998-12-08 Magainin Pharmaceuticals Inc. Certain aminosterol compounds and pharmaceutical compositions including these compounds
US5874597A (en) * 1995-06-07 1999-02-23 Magainin Pharmaceuticals, Inc. Certain aminosterol compounds and pharmaceutical compositions including these compounds
US5792635A (en) * 1995-06-07 1998-08-11 Magainin Pharmaceuticals, Inc. Method of inhibiting the sodium/proton exchanger NHE3 and method of inhibiting growth by administering squalamine
US5840740A (en) * 1995-06-07 1998-11-24 Magainin Pharmaceuticals Inc. Aminosterol compounds and a method of treating infection using the aminosterol compounds
US20060166950A1 (en) * 1995-06-07 2006-07-27 Genaera Corporation Treatment of neovascularization disorders with squalamine
US20090137518A1 (en) * 1995-10-23 2009-05-28 The Children's Medical Center Corporation Therapeutic Antiangiogenic Endostatin Compositions
US20050282253A1 (en) * 1995-10-23 2005-12-22 Folkman M J Therapeutic antiangiogenic endostatin compositions
US7867975B2 (en) 1995-10-23 2011-01-11 The Children's Medical Center Corporation Therapeutic antiangiogenic endostatin compositions
US7495089B2 (en) 1995-10-23 2009-02-24 The Children's Medical Center Corporation Therapeutic antiangiogenic endostatin compositions
US6596712B2 (en) 1996-04-26 2003-07-22 Genaera Corporation Treatment of carcinomas using squalamine in combination with other anti-cancer agents or modalities
US6147060A (en) * 1996-04-26 2000-11-14 Magainin Pharmaceuticals Treatment of carcinomas using squalamine in combination with other anti-cancer agents
US20070049566A1 (en) * 1996-11-05 2007-03-01 D Amato Robert J Methods and Compositions for inhibition of angiogenesis
EP2143444A1 (en) 1996-11-05 2010-01-13 The Children's Medical Center Corporation Compositions for inhibition of anglogenesis
EP2298300A1 (en) 1996-11-05 2011-03-23 The Children's Medical Center Corporation Compositions for inhibition of anglogenesis comprising thalidomide and aspirin
US20090036412A1 (en) * 1996-11-05 2009-02-05 D Amato Robert J Methods and Compositions for Inhibition of Angiogenesis
US7435745B2 (en) 1996-11-05 2008-10-14 Celgene Corporation Methods and compositions for inhibition of angiogenesis
EP1920773A1 (en) 1996-11-05 2008-05-14 The Children's Medical Center Corporation Compositions for inhibition of anglogenesis
US8039488B2 (en) 1996-11-05 2011-10-18 Children's Medical Center Corporation Methods and compositions for inhibition of angiogenesis
EP1586322A2 (en) 1996-11-05 2005-10-19 The Children's Medical Center Corporation Compositions for inhibition of anglogenesis
US20030191098A1 (en) * 1996-11-05 2003-10-09 D'amato Robert J. Methods and compositions for inhibition of angiogenesis
US6262283B1 (en) 1996-12-06 2001-07-17 Magainin Pharmaceuticals Inc. Stereoselective synthesis of 24-hydroxylated compounds useful for the preparation of aminosterols, vitamin D analogs, and other compounds
US6610866B2 (en) 1996-12-06 2003-08-26 Magainin Pharmaceuticals, Inc. Stereoselective synthesis of 24-hydroxylated compounds useful for the preparation of aminosterols, vitamin D analogs, and other compounds
US5981471A (en) * 1997-02-06 1999-11-09 Entremed, Inc. Compositions and methods for inhibiting cellular proliferation
US6734163B2 (en) 1997-02-06 2004-05-11 Entremed, Inc. Compositions and methods for inhibiting cellular proliferation
US6946439B2 (en) 1997-02-06 2005-09-20 Entre Med, Inc. Compositions and methods for inhibiting cellular proliferation comprising TFPI fragments
US20060025329A1 (en) * 1997-02-06 2006-02-02 Todd Hembrough Compositions and methods for inhibiting cellular proliferation comprising TFPI fragments
US20020002294A1 (en) * 1997-09-24 2002-01-03 D' Amato Robert J. Estrogenic compounds as antiangiogenic agents
US6518298B2 (en) 1997-10-16 2003-02-11 Entremed, Inc. Methods and compositions for inhibition of angiogenesis with EM-138
US6228879B1 (en) 1997-10-16 2001-05-08 The Children's Medical Center Methods and compositions for inhibition of angiogenesis
US6150407A (en) * 1998-03-25 2000-11-21 Large Scale Biology Corporation Methods for inhibiting angiogenesis
US6433012B1 (en) * 1998-03-25 2002-08-13 Large Scale Biology Corp. Method for inhibiting inflammatory disease
US20040127545A1 (en) * 1998-05-11 2004-07-01 Childrens' Medical Corporation Analogs of 2-phthalimidinoglutaric acid
US7112602B2 (en) 1998-05-11 2006-09-26 The Children's Medical Center Corporation Methods of treating undesired angiogenesis with 2-methyl-EM-138
US6673828B1 (en) 1998-05-11 2004-01-06 Children's Medical Center Corporation Analogs of 2-Phthalimidinoglutaric acid
US6605622B2 (en) 1998-08-11 2003-08-12 Entremed, Inc. Use of anti-estrogenic compounds as anti-fungal agents
US6420345B1 (en) 1999-03-01 2002-07-16 Cell Genesys, Inc. Methods and reagents for inhibiting angiogenesis
US20040097479A1 (en) * 1999-08-23 2004-05-20 Agoston Gregory E. Methods of obtaining 2-methoxyestradiol of high purity
US7235540B2 (en) 1999-08-23 2007-06-26 Entremed, Inc. Methods of using 2-methoxyestradiol of high purity
US20060025619A1 (en) * 1999-08-23 2006-02-02 Agoston Gregory E Methods of using 2-methoxyestradiol of high purity
US7087592B1 (en) 1999-08-23 2006-08-08 Entre Med, Inc. Compositions comprising purified 2-methoxyestradiol and methods of producing same
US6933383B2 (en) 2000-04-12 2005-08-23 Genaera Corporation Regioselective and stereoselective oxidation of fused ring systems useful for the preparation of aminosterols
US20030171576A1 (en) * 2000-04-12 2003-09-11 Kinney William A. Regioselective and stereoselective oxidation of fused ring systems useful for the preparation of aminosterols
US7728157B2 (en) 2000-04-12 2010-06-01 Ohr Pharmaceutical Inc. Regioselective and stereoselective oxidation of fused ring systems useful for the preparation of aminosterols
US20050187202A1 (en) * 2000-04-12 2005-08-25 Genaera Corporation Regioselective and stereoselective oxidation of fused ring systems useful for the preparation of aminosterols
US20040037875A1 (en) * 2000-07-12 2004-02-26 Metselaar Josbert Maarten Composition for treatment of inflammatory disorders
US6995278B2 (en) 2000-08-18 2006-02-07 Entre Med, Inc. Antiangiogenic agents
US7135581B2 (en) 2000-08-18 2006-11-14 Entremed, Inc. Antiangiogenic agents
US20060135796A1 (en) * 2000-08-18 2006-06-22 Agoston Gregory E Antiangiogenic agents
US20050192258A1 (en) * 2000-08-18 2005-09-01 Agoston Gregory E. Antiangiogenic agents
US7812169B2 (en) 2000-11-30 2010-10-12 The Children's Medical Center Corporation Method of synthesis of 4-amino-thalidomide enantiomers
US20040147558A1 (en) * 2000-11-30 2004-07-29 Anthony Treston Synthesis of 3-amino-thalidomide and its enantiomers
US8153806B2 (en) 2000-11-30 2012-04-10 The Children's Medical Center Corporation Synthesis of 4-amino-thalidomide enantiomers
US20100280249A1 (en) * 2000-11-30 2010-11-04 The Children's Medical Center Corporation Synthesis of 4-amino-thalidomide enantiomers
US20080306113A1 (en) * 2000-11-30 2008-12-11 Anthony Treston Methods for treating macular degeneration using 4-(amino)-2-(2,6-dioxo(3-piperidyle))-isoindoline-1,3-dione
US7955618B2 (en) 2000-12-07 2011-06-07 Enceladus Pharmaceuticals B.V. Composition for treatment of inflammatory disorders
US20030236439A1 (en) * 2002-01-30 2003-12-25 Gregory Agoston Non-steroidal analogs of 2-methoxyestradiol
US20030164219A1 (en) * 2002-02-20 2003-09-04 Joerg Brahm Headliner/duct assembly and welding process therefor
US7977379B2 (en) 2002-05-15 2011-07-12 Bhn Co., Ltd. Method for angiogenesis inhibition or immunostimulation
US20040029955A1 (en) * 2002-05-15 2004-02-12 Takashi Kouge Composition for use in prevention or treatment of vascular-related diseases
EP2316455A1 (en) 2002-05-17 2011-05-04 Celgene Corporation Pharmaceutical compositions for treating cancer
EP2087891A2 (en) 2002-05-17 2009-08-12 Celgene Corporation Pharmaceutical compositions for treating lymphoma
EP2561874A2 (en) 2002-05-17 2013-02-27 Celgene Corporation Pharmaceutical compositions for treating cancer
EP2272512A1 (en) 2002-05-17 2011-01-12 Celgene Corporation Pharmaceutical compositions for treating cancer
EP2258363A1 (en) 2002-05-17 2010-12-08 Celgene Corporation Compositions for treatment of cancers
WO2003097052A2 (en) 2002-05-17 2003-11-27 Celgene Corporation Methods and compositions using immunomodulatory compounds for treatment and management of cancers and other diseases
EP2915533A1 (en) 2002-05-17 2015-09-09 Celgene Corporation Pharmaceutical compositions for treating cancer
EP2272513A1 (en) 2002-05-17 2011-01-12 Celgene Corporation Pharmaceutical compositions for treating cancer
EP2105135A2 (en) 2002-05-17 2009-09-30 Celgene Corporation Pharmaceutical compositions for treating cancer
EP2105136A2 (en) 2002-05-17 2009-09-30 Celgene Corporation Pharmaceutical compositions for treating cancer
US20090226509A1 (en) * 2002-06-12 2009-09-10 Enceladus Pharmaceuticals B.V. Composition for treatment of inflammatory disorders
US20050202078A1 (en) * 2002-08-27 2005-09-15 Schiffelers Raymond M. Vesicle-encapsulated corticosteroids for the treatment of cancer
EP2267031A1 (en) 2003-03-04 2010-12-29 Kirin Beer Kabushiki Kaisha Endothelial cell specific antibodies and uses thereof
EP2264073A1 (en) 2003-03-04 2010-12-22 Kirin Beer Kabushiki Kaisha Endothelial cell specific antibodies and uses thereof
US20070020271A1 (en) * 2003-03-04 2007-01-25 Beverly Teicher Endothelial cell specific antibodies and uses thereof
EP2460522A2 (en) 2003-05-15 2012-06-06 Celgene Corporation Methods and compositions using immunomodulatory compounds for treatment and management of cancers and other diseases
US20050014737A1 (en) * 2003-05-28 2005-01-20 Agoston Gregory E. Antiangiogenic agents
US20070135400A1 (en) * 2003-05-28 2007-06-14 Agoston Gregory E Estradiol derivatives
US7371741B2 (en) 2003-05-28 2008-05-13 Entremed, Inc. Estradiol derivatives and pharmaceutical compositions using same
EP2077112A1 (en) 2003-11-06 2009-07-08 Celgene Corporation Methods and compositions using thalidomide for the treatment and management of cancers and idiopathic pulmonary fibrosis
US8158612B2 (en) 2004-03-12 2012-04-17 Entremed, Inc. Methods of treating disease states using antiangiogenic agents
US7498322B2 (en) 2004-03-12 2009-03-03 Entremed, Inc. Antiangiogenic agents
US20070004689A1 (en) * 2004-03-12 2007-01-04 Agoston Gregory E Antiangiogenic agents
US20050203075A1 (en) * 2004-03-12 2005-09-15 Agoston Gregory E. Antiangiogenic agents
US20060116360A1 (en) * 2004-11-29 2006-06-01 Fogler William E Method of administering anti-angiogenic agents and a method of treating disease using same
US20070111972A1 (en) * 2005-07-12 2007-05-17 Dmi Biosciences, Inc. "methods and products for treatment of diseases"
US8586568B2 (en) 2005-07-12 2013-11-19 Ampio Pharmaceuticals, Inc. Methods and products for treatment of diseases
US20110171306A1 (en) * 2005-07-12 2011-07-14 David Bar-Or Methods and products for treatment of diseases
US20110171307A1 (en) * 2005-07-12 2011-07-14 David Bar-Or Methods and products for treatment of diseases
US8722651B2 (en) 2005-07-12 2014-05-13 Ampio Pharmaceuticals, Inc. Methods and products for treatment of diseases
WO2007027527A2 (en) 2005-08-31 2007-03-08 Celgene Corporation Isoindole-imide compounds and compositions comprising and methods of using the same
US20070185069A1 (en) * 2005-11-14 2007-08-09 Plum Stacy M Anti-angiogenic activity of 2-methoxyestradiol in combination with anti-cancer agents
US20080090760A2 (en) * 2005-12-09 2008-04-17 Todd Hembrough Compositions and Methods for Inhibiting Cellular Proliferation
US20070203066A1 (en) * 2005-12-09 2007-08-30 Holmes Christopher P Compositions and methods for inhibiting cellular proliferation
US20070176403A1 (en) * 2006-02-01 2007-08-02 Dennis Calderone Air adjustable seat
US8399440B2 (en) 2006-03-20 2013-03-19 Entremed, Inc. Disease modifying anti-arthritic activity of 2-methoxyestradiol
US20070231410A1 (en) * 2006-03-20 2007-10-04 Fogler William E Disease modifying anti-arthritic activity of 2-Methoxyestradiol
EP3239144A1 (en) 2006-09-26 2017-11-01 Celgene Corporation 5-substituted quinazolinone derivatives as anti-cancer agents
EP2428513A1 (en) 2006-09-26 2012-03-14 Celgene Corporation 5-substituted quinazolinone derivatives as anti-cancer agents
EP2420497A1 (en) 2006-09-26 2012-02-22 Celgene Corporation 5-substituted quinazolinone derivatives as anti-cancer agents
EP2420498A1 (en) 2006-09-26 2012-02-22 Celgene Corporation 5-substituted quinazolinone derivatives as anti-cancer agents
US20080234243A1 (en) * 2007-01-31 2008-09-25 Lavallee Theresa M Method of treating amyloidosis mediated diseases
EP3101017A1 (en) 2007-03-20 2016-12-07 Celgene Corporation 4'-o-substituted isoindoline derivatives and compositions comprising and methods of using the same
WO2009042177A1 (en) 2007-09-26 2009-04-02 Celgene Corporation 6-, 7-, or 8-substituted quinazolinone derivatives and compositions comprising and methods of using the same
US20090298882A1 (en) * 2008-05-13 2009-12-03 Muller George W Thioxoisoindoline compounds and compositions comprising and methods of using the same
US10206933B2 (en) 2008-05-28 2019-02-19 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-kB for treatment of disease
US8207151B2 (en) 2008-05-28 2012-06-26 Validus Biopharma Inc. Non-hormonal steroid modulators of NF-κB for treatment of disease
US9649320B2 (en) 2008-05-28 2017-05-16 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-κB for treatment of disease
US8334279B2 (en) 2008-05-28 2012-12-18 Validus Genetics Non-hormonal steroid modulators of NF-κB for treatment of disease
US9434758B2 (en) 2008-05-28 2016-09-06 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-κB for treatment of disease
US20100087408A1 (en) * 2008-05-28 2010-04-08 Validus Genetics NON-HORMONAL STEROID MODULATORS OF NF-kB FOR TREATMENT OF DISEASE
US10857161B2 (en) 2008-05-28 2020-12-08 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-kB for treatment of disease
US8673887B2 (en) 2008-05-28 2014-03-18 Reveragen Biopharma, Inc Non-hormonal steroid modulators of NF-kB for treatment of disease
US11833159B2 (en) 2008-05-28 2023-12-05 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-kB for treatment of disease
EP3061758A1 (en) 2008-05-30 2016-08-31 Celgene Corporation 5-substituted isoindoline compounds
EP2749559A1 (en) 2008-05-30 2014-07-02 Celgene Corporation 5-substituted isoindoline compounds
EP3327013A1 (en) 2008-05-30 2018-05-30 Celgene Corporation 5-substituted isoindoline compounds
EP2985281A2 (en) 2008-10-29 2016-02-17 Celgene Corporation Isoindoline compounds for use in the treatment of cancer
WO2010093434A1 (en) 2009-02-11 2010-08-19 Celgene Corporation Isotopologues of lenalidomide
US9611465B2 (en) 2009-05-25 2017-04-04 Celgene Corporation Pharmaceutical composition containing core factor involved in proliferation and differentiation of central nervous cell
US9233113B2 (en) 2009-06-22 2016-01-12 Ampio Pharmaceuticals, Inc. Method for treatment of diseases
US9987292B2 (en) 2009-06-22 2018-06-05 Ampio Pharmaceuticals, Inc. Method for treatment of diseases
US8227457B2 (en) 2009-06-22 2012-07-24 Dmi Acquisition Corp. Method for treatment of diseases
US20100324005A1 (en) * 2009-06-22 2010-12-23 Dmi Acquisition Corp. Method for treatment of diseases
US20100323991A1 (en) * 2009-06-22 2010-12-23 Dmi Acquisition Corp. Methods and products for treatment of diseases
EP3599236A1 (en) 2010-02-11 2020-01-29 Celgene Corporation Arylmethoxy isoindoline derivatives and compositions comprising and methods of using the same
WO2011100380A1 (en) 2010-02-11 2011-08-18 Celgene Corporation Arylmethoxy isoindoline derivatives and compositions comprising and methods of using the same
EP3106460A1 (en) 2010-02-11 2016-12-21 Celgene Corporation Arylmethoxy isoindoline derivatives and compositions comprising and methods of using the same
EP4289838A2 (en) 2010-02-11 2023-12-13 Celgene Corporation Arylmethoxy isoindoline derivatives and compositions comprising and methods of using the same
US10000525B2 (en) 2010-04-05 2018-06-19 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-κB for treatment of disease
US9198921B2 (en) 2010-04-05 2015-12-01 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-κB for treatment of disease
US8802685B2 (en) 2011-03-11 2014-08-12 Celgene Corporation Solid forms of 3-(5-amino-2-methyl-4-oxo-4H-quinazolin-3-yl)-piperidine-2,6-dione, and their pharmaceutical compositions and uses
US9969713B2 (en) 2011-03-11 2018-05-15 Celgene Corporation Solid forms of 3-(5-amino-2-methyl-4-oxo-4H-quinazolin-3-yl)-piperidine-2,6-dione, and their pharmaceutical compositions and uses
WO2012125438A1 (en) 2011-03-11 2012-09-20 Celgene Corporation Solid forms of 3-(5-amino-2methyl-4-oxo-4h-quinazolin-3-yl)-piperidine-2,6-dione, and their pharmaceutical compositions and uses
US9751853B2 (en) 2011-03-11 2017-09-05 Celgene Corporation Solid forms of 3-(5-amino-2-methyl-4-oxo-4H-quinazolin-3-yl)-piperidine-2,6-dione, and their pharmaceutical compositions and uses
US9249121B2 (en) 2011-03-11 2016-02-02 Celgene Corporation Solid forms of 3-(5-amino-2-methyl-4-oxo-4H-quinazolin-3-yl)-piperidine-2,6-dione, and their pharmaceutical compositions and uses
EP3309153A1 (en) 2011-03-11 2018-04-18 Celgene Corporation Solid forms of 3-(5-amino-2-methyl-4-oxo-4h-quinazolin-3-yl)-piperidine-2,6-dione, and their pharmaceutical compositions and uses
US10047151B2 (en) 2011-04-29 2018-08-14 Celgene Corporation Methods for the treatment of cancer and inflammatory diseases using cereblon as a predictor
US9365640B2 (en) 2011-04-29 2016-06-14 Celgene Corporation Methods for the treatment of cancer and inflammatory diseases using cereblon as a predictor
EP3096142A2 (en) 2011-04-29 2016-11-23 Celgene Corporation Methods for the treatment of cancer and inflammatory diseases using cereblon as a predictor
WO2012149299A2 (en) 2011-04-29 2012-11-01 Celgene Corporaiton Methods for the treatment of cancer and inflammatory diseases using cereblon as a predictor
WO2012177678A2 (en) 2011-06-22 2012-12-27 Celgene Corporation Isotopologues of pomalidomide
US9857359B2 (en) 2012-06-29 2018-01-02 Celgene Corporation Methods for determining drug efficacy using cereblon-associated proteins
EP3950681A2 (en) 2012-08-09 2022-02-09 Celgene Corporation Salts and solid forms of the compound (s)-3-(4-((4-morpholinomethyl)benzyl)oxy)-1-oxoisoindolin-2-yl)piperidine-2,6-dione
EP3524598A1 (en) 2012-08-09 2019-08-14 Celgene Corporation A solid form of (s)-3-(4-((4-morpholinomethyl)benzyl)oxy)-1-oxoisoindolin-2-yl)piperidine-2,6-dione hydrochloride
US9587281B2 (en) 2012-08-14 2017-03-07 Celgene Corporation Cereblon isoforms and their use as biomarkers for therapeutic treatment
US10058562B2 (en) 2012-12-19 2018-08-28 Ampio Pharmaceuticals, Inc. Methods of treatment of diseases
US9351979B2 (en) 2012-12-19 2016-05-31 Ampio Pharmaceuticals, Inc. Methods of treatment of diseases
US10092555B2 (en) 2014-06-27 2018-10-09 Celgene Corporation Compositions and methods for inducing conformational changes in cereblon and other E3 ubiquitin ligases
US11419861B2 (en) 2014-06-27 2022-08-23 Celgene Corporation Compositions and methods for inducing conformational changes in cereblon and other E3 ubiquitin ligases
US10668057B2 (en) 2014-06-27 2020-06-02 Celgene Corporation Compositions and methods for inducing conformational changes in cereblon and other E3 ubiquitin ligases
EP3925609A1 (en) 2014-08-22 2021-12-22 Celgene Corporation Methods of treating multiple myeloma with immunomodulatory compounds in combination with antibodies
US10034872B2 (en) 2014-08-22 2018-07-31 Celgene Corporation Methods of treating multiple myeloma with immunomodulatory compounds in combination with antibodies
US11690853B2 (en) 2015-06-29 2023-07-04 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-κβ for treatment of disease
US10799514B2 (en) 2015-06-29 2020-10-13 Reveragen Biopharma, Inc. Non-hormonal steroid modulators of NF-kappa beta for treatment of disease
US11733233B2 (en) 2015-12-28 2023-08-22 Celgene Corporation Compositions and methods for inducing conformational changes in cereblon and other E3 ubiquitin ligases
US10830762B2 (en) 2015-12-28 2020-11-10 Celgene Corporation Compositions and methods for inducing conformational changes in cereblon and other E3 ubiquitin ligases
US10189808B2 (en) 2016-01-08 2019-01-29 Celgene Corporation Solid forms of 2-(4-chlorophenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)methyl)-2,2-difluoroacetamide, and their pharmaceutical compositions and uses
EP3808346A1 (en) 2016-01-08 2021-04-21 Celgene Corporation Antiproliferative compounds for use in the treatment of leukemia
US11365184B2 (en) 2016-01-08 2022-06-21 Celgene Corporation Antiproliferative compounds, and their pharmaceutical compositions and uses
US11401257B2 (en) 2016-01-08 2022-08-02 Celgene Corporation Solid forms of 2-(4-chlorophenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)methyl)-2,2-difluoroacetamide, and their pharmaceutical compositions and uses
US10626101B2 (en) 2016-01-08 2020-04-21 Celgene Corporation Solid forms of 2-(4-chlorophenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)methyl)-2,2-difluoroacetamide, and their pharmaceutical compositions and uses
US10618883B2 (en) 2016-01-08 2020-04-14 Celgene Corporation Antiproliferative compounds, and their pharmaceutical compositions and uses
US10227325B2 (en) 2016-01-08 2019-03-12 Celgene Corporation Antiproliferative compounds, and their pharmaceutical compositions and uses
US9938254B2 (en) 2016-01-08 2018-04-10 Celgene Corporation Antiproliferative compounds, and their pharmaceutical compositions and uses
WO2017120415A1 (en) 2016-01-08 2017-07-13 Celgene Corporation Solid forms of 2-(4-chlorophenyl)-n-((2-2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl) methyl)-2,2-difluoroacetamide, and their pharmaceutical compositions and uses
US11382922B2 (en) 2019-03-07 2022-07-12 Reveragen Biopharma, Inc. Aqueous oral pharmaceutical suspension compositions
US11471471B2 (en) 2019-03-07 2022-10-18 Reveragen Biopharma, Inc. Aqueous oral pharmaceutical suspension compositions

Similar Documents

Publication Publication Date Title
US4994443A (en) Inhibition of angiogenesis
US5001116A (en) Inhibition of angiogenesis
EP0114589B1 (en) Inhibition of angiogenesis
CA1330943C (en) Growth inhibiting agent and the use thereof
JP2806454B2 (en) Angiogenesis inhibitor
EP0398925B1 (en) Growth inhibiting agent and the use thereof
US20070054878A1 (en) Use of hyaluronic acid derivatives for inhibiting inflammatory arthritis
Peterson Effects of megestrol acetate on glucose tolerance and growth hormone secretion in the cat
Li et al. Angiostatic steroids potentiated by sulfated cyclodextrins inhibit corneal neovascularization.
CZ257497A3 (en) Use of hyaluronic acid for treating intersticial cystitis
US4757056A (en) Method for tumor regression in rats, mice and hamsters using hexuronyl hexosaminoglycan-containing compositions
Pitcairn et al. Glucocorticoid and antibiotic effect on experimental gram-negative bacteremic shock
US4966890A (en) Method and composition for arresting angiogenesis and capillary, cell or membrane leakage
US4820693A (en) Method and composition for arresting angiogenesis and capillary, cell or membrane leakage
WO1992008466A1 (en) Heparan sulfate biosynthesis primers
EP2830635A1 (en) Combination treatment comprising sulphated glycosaminoglycans for inducing labor
KR100249555B1 (en) Ocular compositions containing steroids and their use in treating glaucoma
EP0299975B1 (en) Composition for arresting angiogenesis and capillary, cell or membrane leakage
McDONALD et al. Hydrocortisone (compound F) in ophthalmology: Clinical and experimental studies
JPH0455171B2 (en)
Beck et al. Effect of heparin, heparin fragments, and corticosteroids on cerebral endothelial cell growth in vitro and in vivo
Kessler et al. Comparison of Duration of Action of Progesterone and 17-α-Hydroxyprogesterone-17-n-Caproate
JP2000309544A (en) Premature birth or abortion-inhibiting agent, cervical canal maturing inhibitor and inhibitor of hyaluronidase
NZ227639A (en) Composition for inhibiting undesired cell or tissue growth comprising a cyclodextrin derivative and a growth-inhibiting compound
US20060079481A1 (en) Method of treating/preventing mucositis

Legal Events

Date Code Title Description
CC Certificate of correction
FEPP Fee payment procedure

Free format text: PAYOR NUMBER ASSIGNED (ORIGINAL EVENT CODE: ASPN); ENTITY STATUS OF PATENT OWNER: LARGE ENTITY

FPAY Fee payment

Year of fee payment: 4

REMI Maintenance fee reminder mailed
LAPS Lapse for failure to pay maintenance fees
FP Lapsed due to failure to pay maintenance fee

Effective date: 19990219

STCH Information on status: patent discontinuation

Free format text: PATENT EXPIRED DUE TO NONPAYMENT OF MAINTENANCE FEES UNDER 37 CFR 1.362